Registration Dossier

Toxicological information

Toxicity to reproduction

Currently viewing:

Administrative data

Endpoint:
one-generation reproductive toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
27 FEB 2009 - 01 FEB 2011
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study
Cross-reference
Reason / purpose:
reference to same study
Reference
Endpoint:
one-generation reproductive toxicity
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Study period:
27 FEB 2009 - 01 FEB 2011
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study
Justification for type of information:
ANALOGUE APPROACH

1. HYPOTHESIS FOR THE ANALOGUE APPROACH
The structures of the target and source substances are identical and differ only with respect to the ratio of enantiomers where the target substance is a single pure L-isomer and the source substance is an equimolar mixture of L and D isomers.

2. SOURCE AND TARGET CHEMICAL(S) (INCLUDING INFORMATION ON PURITY AND IMPURITIES)
The target substance, L-Citronellyl nitrile, is a mono-constituent substance (EC No. 695-909-8, CAS no. 35931-93-2).
The source substance, DL-Citronellyl nitrile, is a mono-constituent substance (EC No. 257-288-8, CAS no. 51566-62-2).
The source and target substances are both of high purity with a low concentration of impurities.

3. ANALOGUE APPROACH JUSTIFICATION
The read across hypothesis is based on structural similarity where the only difference between target and source molecules is the enantiomeric ratio. In a non-chiral environment the target and source chemicals will have identical properties but in the chiral environment of living organisms the enantiomers may possess different carcinogenicity and teratogenicity (in a chiral environment, stereoisomers might experience selective absorption, protein binding, transport, enzyme interactions and metabolism, receptor interactions, and DNA binding). Therefore, as a precaution for the developmental toxicity endpoint it is suggested that the NOAEL 250 mg/kg bw/day for L-Citronellyl nitrile is used instead of 500 mg/kg bw/day, as it is not known which form is more potent in vivo. All other endpoints are considered to be acceptable for this substance assuming that 50% of the target compound is available in the test material.

4. DATA MATRIX
Please refer to the data matrix included in the read-across justification document attached in Section 13.2.
Reason / purpose:
read-across source
Qualifier:
according to
Guideline:
OECD Guideline 415 [One-Generation Reproduction Toxicity Study (before 9 October 2017)]
Version / remarks:
Deviations see Deviation (other information):
-Acclimation
-Formulation Preparation
-Dosage Administration
-Clinical Observations
-Litter Observations
-Sexual Maturation
-Gross Necropsy, Organ Weights and Tissue Retention
Deviations:
yes
Remarks:
The deviations from the protocol had no impact on the study outcome or interpretation of the data because, relative to the total number of animals evaluated and/or the number of data points collected per parameter, the deviations were not significant.
GLP compliance:
yes
Limit test:
no
Justification for study design:
The requirements of the Organisation for Economic Co-operation and Development Guidelines for Testing of Chemicals No. 415 were used as the basis for study design.
Species:
rat
Strain:
other: Crl:CD(SD)
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Age at study initiation: male: 37 days (age at arrival), female 65 days (age at arrival)
- Weight at study initiation: (P) Males: 129 - 160 g; Females: 224 - 266 g
- Housing: individually housed in stainless steel, wire-bottomed cages; during cohabitation, each pair of male and female rats was housed in the male rat's cage; P generation female rats were individually housed in nesting boxes beginning no later than day of (presumed) gestation 20
- Diet: ad libitum
- Water: ad libitum
ENVIRONMENTAL CONDITIONS
- Temperature: 19°C to 25°C
- Humidity: 30% to 70%
- Air changes: a minimum of ten changes per hour
- Photoperiod: 12-hours light:12-hours dark fluorescent light cycle
- Enrichment: supplied to all rats during the course of the study
Route of administration:
oral: gavage
Vehicle:
corn oil
Details on exposure:
PREPARATION OF DOSING SOLUTIONS:
Results indicate that the formulations are within the acceptable limits of ±15% of nominal concentrations. The formulations are also within the acceptable limits of ≤5% RSD for homogeneity

DIET PREPARATION
Dosages were adjusted weekly for body weight changes and given at approximately the same time each day. Any dams in the process of parturition were not given the test substance and/or vehicle until the following work day. No dam missed more than one daily dosage administration, and such events were noted in the raw data. Prepared formulations were stirred continuously during dosage administration.

VEHICLE
- Justification for use and choice of vehicle (if other than water):
- Concentration in vehicle: 0, 10, 30, 100, or 300 mg/kg/day of test item
- Amount of vehicle: 4 mL/kg for 0 mg/kg/day of test item
Details on mating procedure:
- M/F ratio per cage:one male rat per female rat
- Length of cohabitation: a maximum of 13 days
- Proof of pregnancy: spermatozoa observed in a smear of the vaginal contents and/or a copulatory plug observed in situ
- After successful mating each pregnant female was caged: individual housing
- Any other deviations from standard protocol: see Deviation (other information); cohabitation: one male rat per female rat within each dosage group, except for male rat 777 (Group IV, 500 mg/kg/day), which was cohabitated with a second female due to the unscheduled euthanasia of male rat 796
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Samples of dose formulations were analyzed by HPLC-UV.
The method was validated for the analysis of dose formulations at concentrations ranging from 1.2 mg/mL to 150 mg/mL of the test item in corn oil.
Results for all dose formulations met the acceptance criteria for concentration (within ±15% of nominal concentration) and homogeneity (≤5% relative standard deviation) except for end of study samples for Group II (18.75 mg/mL), Group III (50 mg/mL), Group IV (125 mg/mL) are out of specification (+27.3%, +20.3% and +20.0%, respectively). Time zero stability samples at 125 mg/mL were not within the acceptable limits of ±15% of nominal concentration, (+25.6%); however, the result was accepted and used as the reference concentration for formulation stability evaluations.
Duration of treatment / exposure:
male: once daily beginning 83 days before the cohabitation period, through the cohabitation period (maximum of 13 days) and continuing until the day before euthanasia
female: once daily beginning 14 days before the cohabitation period, through the cohabitation period (maximum of 13 days) and continuing through the day of euthanasia (through day 25 of presumed gestation [rats that do not deliver] or day 22 of lactation [rats that deliver a litter])
F1 generation pups were not directly administered the test substance and/or the vehicle.
Frequency of treatment:
male and female: once daily
Dose / conc.:
0 mg/kg bw/day
Remarks:
dosage volume: 4 mL/kg (adjusted weekly)
Dose / conc.:
75 mg/kg bw/day
Remarks:
dosage volume: 4 mL/kg (adjusted weekly)
Dose / conc.:
200 mg/kg bw/day
Remarks:
dosage volume: 4 mL/kg (adjusted weekly)
Dose / conc.:
500 mg/kg bw/day
Remarks:
dosage volume: 4 mL/kg (adjusted weekly)
No. of animals per sex per dose:
25
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: on the basis of a dosage-range reproduction study
- Rationale for animal assignment: random
- Route administration rationale: The oral (gavage) route was selected for use because: 1) in comparison with the dietary route, the exact dosage can be accurately administered; and 2) it is one possible route of human exposure.
Parental animals: Observations and examinations:
DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: twice each day

BODY WEIGHT: Yes
- Time schedule for examinations: at least weekly during the acclimation period and on the first day of dosage administration, weekly during the dosage period and once on the day of euthanasia (males and females), on DGs 0, 7, 10, 14, 18, 21 and 25 (for rats that did not deliver a litter) and on DLs 1, 5, 8, 11, 15 and 22 (terminal body weight) (females)

FOOD CONSUMPTION AND COMPOUND INTAKE:
- Food consumption: weekly during the acclimation and dosage periods, except during cohabitation. Feed consumption values were also recorded for female rats on DGs 0, 7, 10, 14, 18, 21 and 25 (for rats that did not deliver a litter) and on DLs 1, 5, 8, 11 and 15.Because pups begin to consume maternal feed on or about DL 15, feed consumption values were not tabulated after DL 15.

OTHER:
Female rats were evaluated for adverse clinical signs observed during parturition, duration of gestation (DG 0 to the day the first pup was observed), litter size (all pups delivered) and pup viability at birth. Maternal behavior was evaluated on DLs 1, 5, 8, 15 and 22.
Oestrous cyclicity (parental animals):
Evaluated daily by examination of vaginal cytology
Sperm parameters (parental animals):
Parameters examined in P male parental generations:
Right testis, left testis, left epididymis (whole and cauda), right epididymis, seminal vesicles (with and without fluid) and prostate. Sperm concentration and motility were evaluated.
Litter observations:
STANDARDISATION OF LITTERS
- Performed on day 4 postpartum: no

PARAMETERS EXAMINED
The following parameters were examined in F1 offspring:
number and sex of pups, stillbirths, live births and gross alterations

Postmortem examinations (parental animals):
SACRIFICE
- Male animals: All surviving animals, after completion of the cohabitation period
- Maternal animals: All surviving animals, after completion of the 22-day postpartum period

GROSS NECROPSY
Initial physical examination of external surfaces and all orifices, as well as an internal examination of tissues and organs in situ. The following were examined: external and internal portions of all hollow organs; the external surfaces of the brain and spinal column; the nasal cavity and neck with associated organs and tissues; the thoracic, abdominal and pelvic cavities with associated organs and tissues; and the musculo/skeletal carcass. The lungs were perfused with 10% NBF.

Male rats that died or were euthanized before scheduled termination were examined for the cause of death or condition on the day the observation was made. The rats were examined for gross lesions.

HISTOPATHOLOGY / ORGAN WEIGHTS
The tissues indicated in Table 1 were prepared for microscopic examination and weighed, respectively.
Postmortem examinations (offspring):
SACRIFICE
- The F1 offspring: pups ≤ 14 days of age
- All pups not selected for continued evaluation: day 22 postpartum
- F1 Generation Rats: day 60 (± 3) postpartum

GROSS NECROPSY
- Necropsy included a single cross-section of the head at the level of the frontal-parietal suture and examination of the cross-sectioned brain for apparent hydrocephaly. Carcasses of pups
not selected for continued observation were discarded without further evaluation.

Pups that died before examination of the litter for pup viability were evaluated for vital status at birth. They were examined for gross lesions and the cause of death or condition on the day the observation was made. Pups found on days 2 to 5 postpartum were preserved in Bouin's solution for possible future evaluation; pups found on days 6 to 22 postpartum were preserved in 10% NBF.

- Gross necropsy of the F1 Generation Rats consisted of thoracic, abdominal and pelvic viscera.

HISTOPATHOLOGY / ORGAN WEIGTHS
The tissues indicated in Table 2 were prepared for microscopic examination and weighed, respectively.
Statistics:
All data were tabulated, summarized and/or statistically analyzed:
-body weights, feed consumption values and percent mortality per litter: Parametric test
- litter size or the day a developmental landmark appeared: Nonparametric test
- Clinical observations and other proportion data: Variance Test for Homogeneity of the Binomial Distribution
Reproductive indices:
Fertility index: Number of pregnancies/number of rats that mated
Rat pregnant/rats in cohabitation: Includes only one pregnancy for each rat that impregnated more than one female rat
Offspring viability indices:
Surviving pups/litter: Average number of live pups per litter, including litters with no surviving pups.
Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
Clinical signs in P generation male rats that were attributed to treatment with the test substance included slight, moderate and extreme excess salivation at 500 mg/kg/day. Slight excess salivation was observed as early as DS 3 and persisted until scheduled euthanasia. Slight excess salivation was observed as early as DS 3 and persisted until scheduled euthanasia.
Each of these clinical observations occurred in significantly more (p≤0.01) P generation male rats in comparison to the vehicle control group values. All P generation male rats in the 500 mg/kg/day dosage group had slight excess salivation on one or more occasions, and 20 of 25 P generation male rats in this same dosage group also had moderate excess salivation. In addition, a low incidence of ungroomed coat (N=4; p≤0.01) occurred at 500 mg/kg/day.
All other clinical observations during the precohabitation, gestation and lactation periods were considered unrelated to the test item because: 1) the incidences were not dosage-dependent; and/or 2) the number of rats affected did not differ significantly from the vehicle control group values.
The statistically significant increase (p≤0.01) in the incidence of soft and liquid feces that occurred in the 200 mg/kg/day dosage group during lactation was considered unrelated to
the test item because the increase was independent of dose.
All other clinical observations were considered unrelated to treatment with the test item.
Mortality:
no mortality observed
Description (incidence):
There were no treatment-related deaths at any dosage level tested.
One male rat in the 200 mg/kg/day dosage group was found dead approximately one hour after dosage administration on DS 116. The cause of death for the rat that died was not determined based on the in-life, postmortem and microscopic observations.
All other P generation male rats survived to scheduled euthanasia.
Body weight and weight changes:
effects observed, non-treatment-related
Description (incidence and severity):
The statistically significant increase (p≤0.05) in body weight gains that occurred in P generation male rats in the 500 mg/kg/day dosage group on DSs 57 to 64 was considered unrelated to treatment with the test item because the increase was transient and did not affect the overall body weight gain.
Body weights and body weight gains during the precohabitation, gestation and lactation periods were unaffected by dosages of the test substance as high as 500 mg/kg/day. All
values were comparable among the four dosage groups.
Statistically significant increases (p≤0.01) in body weight gain occurred at 75 and 500 mg/kg/day on days 1 to 5 of lactation (days of lactation 1 to 5) and at 500 mg/kg/day on days of lactation 1 to 22, in comparison to the vehicle control group values.
The average maternal body weight on day of lactation 22 was significantly increased (p≤0.01) in the 500 mg/kg/day dosage group, as compared to the vehicle control group value. These increases in body weight and body weight gain were considered unrelated to treatment because: 1) the increase was independent of dose; 2) the increase was transient; and/or 3) the increase reflected a net loss in body weight that occurred in the vehicle control group on days of lactation 1 to 5. At 75 mg/kg/day, body weight gains were significantly reduced (p≤0.05) on days of lactation 8 to 11, in comparison to the vehicle control group value. This reduction was considered unrelated to the test item because the reduction was transient and independent of dose.
Terminal body weights for P generation male rats treated with 500 mg/kg/day of the test item were slightly reduced (by 6%), in comparison to the vehicle control group value.
This reduction did not reach statistical significance, but reflected an overall reduction (significant at p≤0.01) in body weight gains that occurred at 500 mg/kg/day for the cumulative dosage period (DSs 1 to 127).
Terminal body weights for P generation female rats treated with 500 mg/kg/day of the test item were significantly increased (p≤0.01), in comparison to the vehicle control group value.
Food consumption and compound intake (if feeding study):
effects observed, non-treatment-related
Description (incidence and severity):
Absolute and relative feed consumption values were unaffected by dosages of the test substance as high as 500 mg/kg/day. All values were comparable among the four dosage groups. Transient, but statistically significant increases (p≤0.05 or p≤0.01) in relative feed consumption occurred at 500 mg/kg/day on DSs 57 to 64, DSs 113 to 120 and DSs 120 to 127, in comparison to the vehicle control group values. These increases were considered unrelated to treatment with the test item because: 1) the increases were transient; and 2) there was no corresponding change in absolute feed consumption in this dosage group.
Organ weight findings including organ / body weight ratios:
effects observed, non-treatment-related
Histopathological findings: non-neoplastic:
effects observed, non-treatment-related
Description (incidence and severity):
There were no gross or microscopic test substance-related pathology findings observed in the rats evaluated. The changes observed in these rats were considered to be incidental or
spontaneous changes commonly observed in control Crl:CD (SD) rats. There were no adverse pathology findings in these rats related to the daily oral gavage of the test substance under the conditions of this study.
Other effects:
no effects observed
Description (incidence and severity):
There were no test substance-related necropsy observations.
Reproductive function: oestrous cycle:
no effects observed
Description (incidence and severity):
The number of estrous stages per 14 days was comparable among the four dosage groups during the precohabitation period.
A slight increase was noted for the mean number of primordial follicles in the 500 mg/kg/day, as compared to the vehicle control group (159 and 132 follicles, respectively). This increase was associated with higher variability than noted for the vehicle control group rats. As well, two rats in the 500 mg/kg/day dosage group (nos. 16289 and 16291) showed comparatively high numbers of primordial follicles (451 and 359 total primordial follicles per rat, respectively), most likely influencing both the noted increase from the vehicle control group rats, and the variability as well. Corpora lutea were present for all animals.
Reproductive function: sperm measures:
no effects observed
Description (incidence and severity):
All sperm parameters evaluated were unaffected by dosages of the test substance as high as 500 mg/kg/day. Values for number and percent motile sperm, number of nonmotile
sperm and total sperm count from the vas deferens and cauda epididymal sperm count and density were comparable among the four dosage groups and did not significantly differ from the vehicle control group values.
Reproductive performance:
no effects observed
Description (incidence and severity):
All mating and fertility parameters [numbers of days in cohabitation, rats that mated, the fertility index (number of pregnancies per number of rats that mated), rats with confirmed
mating dates during the first or second weeks of cohabitation and number of pregnancies per number of rats in cohabitation] were unaffected by dosages of the test substance as
high as 500 mg/kg/day. All values were comparable among the four dosage groups and did not significantly differ from the vehicle control group values.
An overall reduction (significant at p≤0.01) in body weight gains that occurred at 500 mg/kg/day for the cumulative dosage period (DSs 1 to 127).
Organs with statistically significant potentially test item-related weight changes included the brain, liver, kidneys and spleen.
The number of estrous stages per 14 days was comparable among the four dosage groups during the precohabitation period.
Natural delivery and litter observations were unaffected by dosages of the test item as high as 500 mg/kg/day.
The ovarian primordial follicle counts did not appear to be affected by treatment with the test substance at any dosage level tested.
Increased incidences of excess salivation and/or ungroomed coat occurred in P generation male and/or female rats in the 500 mg/kg/day dosage group. However, these clinical signs were not considered an adverse effect of the test item. At 500 mg/kg/day, sporadic reductions in body weight gains occurred in male rats prior to cohabitation, followed by persistent reductions in weight gain through the end of the study. Non-reproductive organ weights were affected in both sexes at the end of the dosage period, as well as terminal body weights. Increased brain and spleen weights occurred in male rats treated with 500 mg/kg/day
test item, while increased liver and kidney weights occurred in both sexes in this same dosage group. The toxicological significance of the increased organ weights was unable to be determined because there were no microscopic findings that could be correlated with the changes in organ weights.
Key result
Dose descriptor:
NOAEL
Remarks:
reproductive
Effect level:
> 500 mg/kg bw/day
Based on:
test mat.
Sex:
male/female
Basis for effect level:
clinical signs
body weight and weight gain
organ weights and organ / body weight ratios
Clinical signs:
effects observed, non-treatment-related
Description (incidence and severity):
Preweaning:
There were no treatment-related clinical signs observed in the F1 generation pups following treatment of P generation rats with the test item at dosages as high as 500 mg/kg/day. All transient and persistent clinical observations were considered unrelated to maternal treatment because: 1) the incidences were not dosage-dependent; 2) several of the observations were presumed related to the tattooing process that occurred on day 1 postpartum; and/or 3) the number of litters
affected did not differ significantly from the vehicle control group values.
Mortality / viability:
mortality observed, non-treatment-related
Description (incidence and severity):
The statistically significant increase (p≤0.01) in the number of pups with an unknown vital status and the significant increase in the number that died between days 16 and 22 postpartum that occurred in the 200 mg/kg/day dosage group was considered unrelated to maternal treatment because these increases were independent of dose.
The significant reduction (p≤0.05) in the viability index that occurred at 500 mg/kg/day was not considered to be adverse because: 1) the value (94.7%) was within the historical range of the Testing Facility (mean: 97.4%; range: 93.8% to 100%); and 2) five of the 13 deaths that occurred in the 500 mg/kg/day dosage group between days 2 and 5 postpartum were in one litter (no. 16281). These increases were considered unrelated to treatment because: 1) the increase was independent of dose; and/or 2) the value was within the historical range of the Testing Facility.
Postweaning:
All clinical observations that occurred in male or female rats during the postweaning period were considered unrelated to treatment of P generation rats with the test item because: 1) the incidences were not dosage-dependent; 2) the observations occurred in only one rat in any dosage group; and/or 3) the observations occurred only in the vehicle control group. These clinical observations included a scab, abrasion and/or ulceration on one or more areas of the body, soft or liquid feces, chromodacryorrhea, abdominal distention, short digits on the left hindpaw, thin or sparse hair coat, localized alopecia (neck), bent tail, misaligned/missing/broken incisors, mild dehydration (based on skin turgor), red substance on the fur, ungroomed coat and a swollen snout.
Body weight and weight changes:
effects observed, non-treatment-related
Description (incidence and severity):
Body weights and body weight gains of the F1 generation male and female rats during the postweaning period were unaffected by treatment of P generation rats with the test item at dosages as high as 500 mg/kg/day.
The statistically significant increase (p≤0.05 or p≤0.01) in body weight gains that occurred in F1 generation female rats at 75 and 200 mg/kg/day on days 51 to 57 postpartum was not attributed to maternal treatment because: 1) the increase was transient; and 2) the increase was independent of dose.
Food consumption and compound intake (if feeding study):
effects observed, non-treatment-related
Description (incidence and severity):
Absolute and relative feed consumption values for F1 generation male and female rats during the postweaning period were unaffected by treatment of P generation rats with the test item at dosages as high as 500 mg/kg/day.
Absolute feed consumption values for F1 male rats in the 75, 200 and 500 mg/kg/day dosage groups were 103%, 100% and 101% of the vehicle control group value, respectively, on days 23 to 57 postpartum. Absolute feed consumption values for F1 female rats in the 75, 200 and 500 mg/kg/day dosage groups were 101%, 102% and 104% of the vehicle control group value, respectively, on days 23 to 57 postpartum. The statistically significant reductions (p≤0.05) in relative feed consumption that occurred in female rats at 75 mg/kg/day (days 23 to 30 postpartum) and in male rats at 200 mg/kg/day (day 37 to 44 postpartum) were not attributed to maternal treatment because the reductions were transient and independent of dose.
Sexual maturation:
no effects observed
Description (incidence and severity):
Sexual maturation in F1 rats was unaffected by treatment of P generation rats with the test substance at dosages as high as 500 mg/kg/day.
The average day on which vaginal opening was observed in F1 female rats was comparable among the dosage groups. In addition, the average body weight on the day sexual maturation was achieved in F1 male and female rats was comparable among the dosage groups and did not differ significantly from the vehicle control group values.
Organ weight findings including organ / body weight ratios:
effects observed, non-treatment-related
Description (incidence and severity):
There were no apparent effects of treatment of P generation rats on the reproductive and nonreproductive tissue weights of the F1 male or female rats at any dosage level tested.
A slight, but statistically significant increase (p≤0.01) in the absolute weight of the left epididymis occurred in F1 male rats at 500 mg/kg/day, in comparison to the vehicle control group value. This increase was considered unrelated to treatment of P generation rats because there was no statistically significant change in the contralateral organ or in the relative weight of the left epididymis. In addition, the statistically significant reduction (p≤0.01) in the relative (% terminal body weight) weight of the left kidney for F1 male rats at 75 mg/kg/day was not attributed to exposure to the test item because: 1) the reduction was independent of dose; and 2) there was no statistically significant change in the contralateral organ.
Gross pathological findings:
effects observed, non-treatment-related
Description (incidence and severity):
Preweaning:
There were no gross lesions observed in the F1 generation pups that survived to scheduled necropsy on day 22 postpartum. In the pups that were stillborn, found dead or humanely euthanized, no milk was present in the stomach of 5, 1, 1 and 1 F1 generation pups in the 0 (Vehicle), 75, 200 and 500 mg/kg/day dosage groups, respectively.
Postweaning:
There were no test substance-related necropsy observations. All necropsy observations were considered unrelated to treatment of P generation rats because: 1) the incidences were not dosage-dependent; and/or 2) the observations occurred in only one rat in any dosage group. In F1 male rats, these necropsy observations included numerous red areas on the thymus or left lateral lobe of the lung, a mottled (red and dark red) appearance to the right diaphragmatic lobe of the lung, an extra lobe present on the left lateral lobe of the liver, an enlarged right testis, slight dilation of the pelvis in the right kidney, a pale appearance to the left kidney and a clear fluid-filled cyst on the left kidney. Gross lesions observed in F1 female rats included numerous red areas on the thymus, a dark red area on the right diaphragmatic lobe of the lungs and slight dilation of the pelvis in the right kidney. No other gross lesions
occurred.
Other effects:
effects observed, non-treatment-related
Description (incidence and severity):
The lactation index was significantly increased (p≤0.05 or p≤0.01) in the 75 and 500 mg/kg/day dosage groups, in comparison to the vehicle control group value.
This increase was considered unrelated to treatment because: 1) the increase was independent ofdose; and/or 2) the value was within the historical range of the Testing Facility.
Anogenital distance on days 1 or 22 postpartum in F1 male and female pups was not affected by treatment of P generation rats with the test item at any dosage level tested.
Nipple eruption did not occur in any male pup at any dosage level tested. All female pups had nipples present on day 12 postpartum, with the exception of two from litter 16228 (F1 generation; 75 mg/kg/day dosage group).
Key result
Dose descriptor:
NOAEL
Remarks:
viability and growth
Generation:
F1
Effect level:
> 500 mg/kg bw/day
Based on:
test mat.
Sex:
male/female
Basis for effect level:
viability
body weight and weight gain
Key result
Reproductive effects observed:
no
Lowest effective dose / conc.:
500 mg/kg bw/day
Treatment related:
no
Conclusions:
Based on the results of this study, the no-observable-adverse-effect-level (NOAEL) for toxicity of the test item is 200 mg/kg/day.
The reproductive NOAEL in the P generation rats and the NOAEL for viability and growth of the F1 generation offspring is greater than 500 mg/kg/day. There were no apparent effects on estrous cycling, mating and fertility, reproductive organ weights or natural delivery parameters in the P generation and growth and development (including anogenital distance, nipple eruption or sexual maturation) in the F1 generation rats at the highest dosage level tested (500 mg/kg/day).

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2011
Report Date:
2011

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 415 [One-Generation Reproduction Toxicity Study (before 9 October 2017)]
Version / remarks:
Deviations see Deviation (other information):
-Acclimation
-Formulation Preparation
-Dosage Administration
-Clinical Observations
-Litter Observations
-Sexual Maturation
-Gross Necropsy, Organ Weights and Tissue Retention
Deviations:
yes
Remarks:
The deviations from the protocol had no impact on the study outcome or interpretation of the data because, relative to the total number of animals evaluated and/or the number of data points collected per parameter, the deviations were not significant.
GLP compliance:
yes
Limit test:
no
Justification for study design:
The requirements of the Organisation for Economic Co-operation and Development Guidelines for Testing of Chemicals No. 415 were used as the basis for study design.

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
liquid
Details on test material:
Lot (1):
Appearance: Clear colourless to yellowish liquid
Storage: room temperature, protected from light

Lot (2):
Appearance: Clear colourless to yellowish liquid
Storage: room temperature, protected from light

Test animals

Species:
rat
Strain:
other: Crl:CD(SD)
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Age at study initiation: male: 37 days (age at arrival), female 65 days (age at arrival)
- Weight at study initiation: (P) Males: 129 - 160 g; Females: 224 - 266 g
- Housing: individually housed in stainless steel, wire-bottomed cages; during cohabitation, each pair of male and female rats was housed in the male rat's cage; P generation female rats were individually housed in nesting boxes beginning no later than day of (presumed) gestation 20
- Diet: ad libitum
- Water: ad libitum
ENVIRONMENTAL CONDITIONS
- Temperature: 19°C to 25°C
- Humidity: 30% to 70%
- Air changes: a minimum of ten changes per hour
- Photoperiod: 12-hours light:12-hours dark fluorescent light cycle
- Enrichment: supplied to all rats during the course of the study

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
corn oil
Details on exposure:
PREPARATION OF DOSING SOLUTIONS:
Results indicate that the formulations are within the acceptable limits of ±15% of nominal concentrations. The formulations are also within the acceptable limits of ≤5% RSD for homogeneity

DIET PREPARATION
Dosages were adjusted weekly for body weight changes and given at approximately the same time each day. Any dams in the process of parturition were not given the test substance and/or vehicle until the following work day. No dam missed more than one daily dosage administration, and such events were noted in the raw data. Prepared formulations were stirred continuously during dosage administration.

VEHICLE
- Justification for use and choice of vehicle (if other than water):
- Concentration in vehicle: 0, 10, 30, 100, or 300 mg/kg/day of test item
- Amount of vehicle: 4 mL/kg for 0 mg/kg/day of test item
Details on mating procedure:
- M/F ratio per cage:one male rat per female rat
- Length of cohabitation: a maximum of 13 days
- Proof of pregnancy: spermatozoa observed in a smear of the vaginal contents and/or a copulatory plug observed in situ
- After successful mating each pregnant female was caged: individual housing
- Any other deviations from standard protocol: see Deviation (other information); cohabitation: one male rat per female rat within each dosage group, except for male rat 777 (Group IV, 500 mg/kg/day), which was cohabitated with a second female due to the unscheduled euthanasia of male rat 796
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Samples of dose formulations were analyzed by HPLC-UV.
The method was validated for the analysis of dose formulations at concentrations ranging from 1.2 mg/mL to 150 mg/mL of the test item in corn oil.
Results for all dose formulations met the acceptance criteria for concentration (within ±15% of nominal concentration) and homogeneity (≤5% relative standard deviation) except for end of study samples for Group II (18.75 mg/mL), Group III (50 mg/mL), Group IV (125 mg/mL) are out of specification (+27.3%, +20.3% and +20.0%, respectively). Time zero stability samples at 125 mg/mL were not within the acceptable limits of ±15% of nominal concentration, (+25.6%); however, the result was accepted and used as the reference concentration for formulation stability evaluations.
Duration of treatment / exposure:
male: once daily beginning 83 days before the cohabitation period, through the cohabitation period (maximum of 13 days) and continuing until the day before euthanasia
female: once daily beginning 14 days before the cohabitation period, through the cohabitation period (maximum of 13 days) and continuing through the day of euthanasia (through day 25 of presumed gestation [rats that do not deliver] or day 22 of lactation [rats that deliver a litter])
F1 generation pups were not directly administered the test substance and/or the vehicle.
Frequency of treatment:
male and female: once daily
Doses / concentrationsopen allclose all
Dose / conc.:
0 mg/kg bw/day
Remarks:
dosage volume: 4 mL/kg (adjusted weekly)
Dose / conc.:
75 mg/kg bw/day
Remarks:
dosage volume: 4 mL/kg (adjusted weekly)
Dose / conc.:
200 mg/kg bw/day
Remarks:
dosage volume: 4 mL/kg (adjusted weekly)
Dose / conc.:
500 mg/kg bw/day
Remarks:
dosage volume: 4 mL/kg (adjusted weekly)
No. of animals per sex per dose:
25
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: on the basis of a dosage-range reproduction study
- Rationale for animal assignment: random
- Route administration rationale: The oral (gavage) route was selected for use because: 1) in comparison with the dietary route, the exact dosage can be accurately administered; and 2) it is one possible route of human exposure.

Examinations

Parental animals: Observations and examinations:
DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: twice each day

BODY WEIGHT: Yes
- Time schedule for examinations: at least weekly during the acclimation period and on the first day of dosage administration, weekly during the dosage period and once on the day of euthanasia (males and females), on DGs 0, 7, 10, 14, 18, 21 and 25 (for rats that did not deliver a litter) and on DLs 1, 5, 8, 11, 15 and 22 (terminal body weight) (females)

FOOD CONSUMPTION AND COMPOUND INTAKE:
- Food consumption: weekly during the acclimation and dosage periods, except during cohabitation. Feed consumption values were also recorded for female rats on DGs 0, 7, 10, 14, 18, 21 and 25 (for rats that did not deliver a litter) and on DLs 1, 5, 8, 11 and 15.Because pups begin to consume maternal feed on or about DL 15, feed consumption values were not tabulated after DL 15.

OTHER:
Female rats were evaluated for adverse clinical signs observed during parturition, duration of gestation (DG 0 to the day the first pup was observed), litter size (all pups delivered) and pup viability at birth. Maternal behavior was evaluated on DLs 1, 5, 8, 15 and 22.
Oestrous cyclicity (parental animals):
Evaluated daily by examination of vaginal cytology
Sperm parameters (parental animals):
Parameters examined in P male parental generations:
Right testis, left testis, left epididymis (whole and cauda), right epididymis, seminal vesicles (with and without fluid) and prostate. Sperm concentration and motility were evaluated.
Litter observations:
STANDARDISATION OF LITTERS
- Performed on day 4 postpartum: no

PARAMETERS EXAMINED
The following parameters were examined in F1 offspring:
number and sex of pups, stillbirths, live births and gross alterations

Postmortem examinations (parental animals):
SACRIFICE
- Male animals: All surviving animals, after completion of the cohabitation period
- Maternal animals: All surviving animals, after completion of the 22-day postpartum period

GROSS NECROPSY
Initial physical examination of external surfaces and all orifices, as well as an internal examination of tissues and organs in situ. The following were examined: external and internal portions of all hollow organs; the external surfaces of the brain and spinal column; the nasal cavity and neck with associated organs and tissues; the thoracic, abdominal and pelvic cavities with associated organs and tissues; and the musculo/skeletal carcass. The lungs were perfused with 10% NBF.

Male rats that died or were euthanized before scheduled termination were examined for the cause of death or condition on the day the observation was made. The rats were examined for gross lesions.

HISTOPATHOLOGY / ORGAN WEIGHTS
The tissues indicated in Table 1 were prepared for microscopic examination and weighed, respectively.
Postmortem examinations (offspring):
SACRIFICE
- The F1 offspring: pups ≤ 14 days of age
- All pups not selected for continued evaluation: day 22 postpartum
- F1 Generation Rats: day 60 (± 3) postpartum

GROSS NECROPSY
- Necropsy included a single cross-section of the head at the level of the frontal-parietal suture and examination of the cross-sectioned brain for apparent hydrocephaly. Carcasses of pups
not selected for continued observation were discarded without further evaluation.

Pups that died before examination of the litter for pup viability were evaluated for vital status at birth. They were examined for gross lesions and the cause of death or condition on the day the observation was made. Pups found on days 2 to 5 postpartum were preserved in Bouin's solution for possible future evaluation; pups found on days 6 to 22 postpartum were preserved in 10% NBF.

- Gross necropsy of the F1 Generation Rats consisted of thoracic, abdominal and pelvic viscera.

HISTOPATHOLOGY / ORGAN WEIGTHS
The tissues indicated in Table 2 were prepared for microscopic examination and weighed, respectively.
Statistics:
All data were tabulated, summarized and/or statistically analyzed:
-body weights, feed consumption values and percent mortality per litter: Parametric test
- litter size or the day a developmental landmark appeared: Nonparametric test
- Clinical observations and other proportion data: Variance Test for Homogeneity of the Binomial Distribution
Reproductive indices:
Fertility index: Number of pregnancies/number of rats that mated
Rat pregnant/rats in cohabitation: Includes only one pregnancy for each rat that impregnated more than one female rat
Offspring viability indices:
Surviving pups/litter: Average number of live pups per litter, including litters with no surviving pups.

Results and discussion

Results: P0 (first parental generation)

General toxicity (P0)

Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
Clinical signs in P generation male rats that were attributed to treatment with the test substance included slight, moderate and extreme excess salivation at 500 mg/kg/day. Slight excess salivation was observed as early as DS 3 and persisted until scheduled euthanasia. Slight excess salivation was observed as early as DS 3 and persisted until scheduled euthanasia.
Each of these clinical observations occurred in significantly more (p≤0.01) P generation male rats in comparison to the vehicle control group values. All P generation male rats in the 500 mg/kg/day dosage group had slight excess salivation on one or more occasions, and 20 of 25 P generation male rats in this same dosage group also had moderate excess salivation. In addition, a low incidence of ungroomed coat (N=4; p≤0.01) occurred at 500 mg/kg/day.
All other clinical observations during the precohabitation, gestation and lactation periods were considered unrelated to the test item because: 1) the incidences were not dosage-dependent; and/or 2) the number of rats affected did not differ significantly from the vehicle control group values.
The statistically significant increase (p≤0.01) in the incidence of soft and liquid feces that occurred in the 200 mg/kg/day dosage group during lactation was considered unrelated to
the test item because the increase was independent of dose.
All other clinical observations were considered unrelated to treatment with the test item.
Mortality:
no mortality observed
Description (incidence):
There were no treatment-related deaths at any dosage level tested.
One male rat in the 200 mg/kg/day dosage group was found dead approximately one hour after dosage administration on DS 116. The cause of death for the rat that died was not determined based on the in-life, postmortem and microscopic observations.
All other P generation male rats survived to scheduled euthanasia.
Body weight and weight changes:
effects observed, non-treatment-related
Description (incidence and severity):
The statistically significant increase (p≤0.05) in body weight gains that occurred in P generation male rats in the 500 mg/kg/day dosage group on DSs 57 to 64 was considered unrelated to treatment with the test item because the increase was transient and did not affect the overall body weight gain.
Body weights and body weight gains during the precohabitation, gestation and lactation periods were unaffected by dosages of the test substance as high as 500 mg/kg/day. All
values were comparable among the four dosage groups.
Statistically significant increases (p≤0.01) in body weight gain occurred at 75 and 500 mg/kg/day on days 1 to 5 of lactation (days of lactation 1 to 5) and at 500 mg/kg/day on days of lactation 1 to 22, in comparison to the vehicle control group values.
The average maternal body weight on day of lactation 22 was significantly increased (p≤0.01) in the 500 mg/kg/day dosage group, as compared to the vehicle control group value. These increases in body weight and body weight gain were considered unrelated to treatment because: 1) the increase was independent of dose; 2) the increase was transient; and/or 3) the increase reflected a net loss in body weight that occurred in the vehicle control group on days of lactation 1 to 5. At 75 mg/kg/day, body weight gains were significantly reduced (p≤0.05) on days of lactation 8 to 11, in comparison to the vehicle control group value. This reduction was considered unrelated to the test item because the reduction was transient and independent of dose.
Terminal body weights for P generation male rats treated with 500 mg/kg/day of the test item were slightly reduced (by 6%), in comparison to the vehicle control group value.
This reduction did not reach statistical significance, but reflected an overall reduction (significant at p≤0.01) in body weight gains that occurred at 500 mg/kg/day for the cumulative dosage period (DSs 1 to 127).
Terminal body weights for P generation female rats treated with 500 mg/kg/day of the test item were significantly increased (p≤0.01), in comparison to the vehicle control group value.
Food consumption and compound intake (if feeding study):
effects observed, non-treatment-related
Description (incidence and severity):
Absolute and relative feed consumption values were unaffected by dosages of the test substance as high as 500 mg/kg/day. All values were comparable among the four dosage groups. Transient, but statistically significant increases (p≤0.05 or p≤0.01) in relative feed consumption occurred at 500 mg/kg/day on DSs 57 to 64, DSs 113 to 120 and DSs 120 to 127, in comparison to the vehicle control group values. These increases were considered unrelated to treatment with the test item because: 1) the increases were transient; and 2) there was no corresponding change in absolute feed consumption in this dosage group.
Organ weight findings including organ / body weight ratios:
effects observed, non-treatment-related
Histopathological findings: non-neoplastic:
effects observed, non-treatment-related
Description (incidence and severity):
There were no gross or microscopic test substance-related pathology findings observed in the rats evaluated. The changes observed in these rats were considered to be incidental or
spontaneous changes commonly observed in control Crl:CD (SD) rats. There were no adverse pathology findings in these rats related to the daily oral gavage of the test substance under the conditions of this study.
Other effects:
no effects observed
Description (incidence and severity):
There were no test substance-related necropsy observations.

Reproductive function / performance (P0)

Reproductive function: oestrous cycle:
no effects observed
Description (incidence and severity):
The number of estrous stages per 14 days was comparable among the four dosage groups during the precohabitation period.
A slight increase was noted for the mean number of primordial follicles in the 500 mg/kg/day, as compared to the vehicle control group (159 and 132 follicles, respectively). This increase was associated with higher variability than noted for the vehicle control group rats. As well, two rats in the 500 mg/kg/day dosage group (nos. 16289 and 16291) showed comparatively high numbers of primordial follicles (451 and 359 total primordial follicles per rat, respectively), most likely influencing both the noted increase from the vehicle control group rats, and the variability as well. Corpora lutea were present for all animals.
Reproductive function: sperm measures:
no effects observed
Description (incidence and severity):
All sperm parameters evaluated were unaffected by dosages of the test substance as high as 500 mg/kg/day. Values for number and percent motile sperm, number of nonmotile
sperm and total sperm count from the vas deferens and cauda epididymal sperm count and density were comparable among the four dosage groups and did not significantly differ from the vehicle control group values.
Reproductive performance:
no effects observed
Description (incidence and severity):
All mating and fertility parameters [numbers of days in cohabitation, rats that mated, the fertility index (number of pregnancies per number of rats that mated), rats with confirmed
mating dates during the first or second weeks of cohabitation and number of pregnancies per number of rats in cohabitation] were unaffected by dosages of the test substance as
high as 500 mg/kg/day. All values were comparable among the four dosage groups and did not significantly differ from the vehicle control group values.

Details on results (P0)

An overall reduction (significant at p≤0.01) in body weight gains that occurred at 500 mg/kg/day for the cumulative dosage period (DSs 1 to 127).
Organs with statistically significant potentially test item-related weight changes included the brain, liver, kidneys and spleen.
The number of estrous stages per 14 days was comparable among the four dosage groups during the precohabitation period.
Natural delivery and litter observations were unaffected by dosages of the test item as high as 500 mg/kg/day.
The ovarian primordial follicle counts did not appear to be affected by treatment with the test substance at any dosage level tested.
Increased incidences of excess salivation and/or ungroomed coat occurred in P generation male and/or female rats in the 500 mg/kg/day dosage group. However, these clinical signs were not considered an adverse effect of the test item. At 500 mg/kg/day, sporadic reductions in body weight gains occurred in male rats prior to cohabitation, followed by persistent reductions in weight gain through the end of the study. Non-reproductive organ weights were affected in both sexes at the end of the dosage period, as well as terminal body weights. Increased brain and spleen weights occurred in male rats treated with 500 mg/kg/day
test item, while increased liver and kidney weights occurred in both sexes in this same dosage group. The toxicological significance of the increased organ weights was unable to be determined because there were no microscopic findings that could be correlated with the changes in organ weights.

Effect levels (P0)

Key result
Dose descriptor:
NOAEL
Remarks:
reproductive
Effect level:
> 500 mg/kg bw/day
Based on:
test mat.
Sex:
male/female
Basis for effect level:
clinical signs
body weight and weight gain
organ weights and organ / body weight ratios

Results: F1 generation

General toxicity (F1)

Clinical signs:
effects observed, non-treatment-related
Description (incidence and severity):
Preweaning:
There were no treatment-related clinical signs observed in the F1 generation pups following treatment of P generation rats with the test item at dosages as high as 500 mg/kg/day. All transient and persistent clinical observations were considered unrelated to maternal treatment because: 1) the incidences were not dosage-dependent; 2) several of the observations were presumed related to the tattooing process that occurred on day 1 postpartum; and/or 3) the number of litters
affected did not differ significantly from the vehicle control group values.
Mortality / viability:
mortality observed, non-treatment-related
Description (incidence and severity):
The statistically significant increase (p≤0.01) in the number of pups with an unknown vital status and the significant increase in the number that died between days 16 and 22 postpartum that occurred in the 200 mg/kg/day dosage group was considered unrelated to maternal treatment because these increases were independent of dose.
The significant reduction (p≤0.05) in the viability index that occurred at 500 mg/kg/day was not considered to be adverse because: 1) the value (94.7%) was within the historical range of the Testing Facility (mean: 97.4%; range: 93.8% to 100%); and 2) five of the 13 deaths that occurred in the 500 mg/kg/day dosage group between days 2 and 5 postpartum were in one litter (no. 16281). These increases were considered unrelated to treatment because: 1) the increase was independent of dose; and/or 2) the value was within the historical range of the Testing Facility.
Postweaning:
All clinical observations that occurred in male or female rats during the postweaning period were considered unrelated to treatment of P generation rats with the test item because: 1) the incidences were not dosage-dependent; 2) the observations occurred in only one rat in any dosage group; and/or 3) the observations occurred only in the vehicle control group. These clinical observations included a scab, abrasion and/or ulceration on one or more areas of the body, soft or liquid feces, chromodacryorrhea, abdominal distention, short digits on the left hindpaw, thin or sparse hair coat, localized alopecia (neck), bent tail, misaligned/missing/broken incisors, mild dehydration (based on skin turgor), red substance on the fur, ungroomed coat and a swollen snout.
Body weight and weight changes:
effects observed, non-treatment-related
Description (incidence and severity):
Body weights and body weight gains of the F1 generation male and female rats during the postweaning period were unaffected by treatment of P generation rats with the test item at dosages as high as 500 mg/kg/day.
The statistically significant increase (p≤0.05 or p≤0.01) in body weight gains that occurred in F1 generation female rats at 75 and 200 mg/kg/day on days 51 to 57 postpartum was not attributed to maternal treatment because: 1) the increase was transient; and 2) the increase was independent of dose.
Food consumption and compound intake (if feeding study):
effects observed, non-treatment-related
Description (incidence and severity):
Absolute and relative feed consumption values for F1 generation male and female rats during the postweaning period were unaffected by treatment of P generation rats with the test item at dosages as high as 500 mg/kg/day.
Absolute feed consumption values for F1 male rats in the 75, 200 and 500 mg/kg/day dosage groups were 103%, 100% and 101% of the vehicle control group value, respectively, on days 23 to 57 postpartum. Absolute feed consumption values for F1 female rats in the 75, 200 and 500 mg/kg/day dosage groups were 101%, 102% and 104% of the vehicle control group value, respectively, on days 23 to 57 postpartum. The statistically significant reductions (p≤0.05) in relative feed consumption that occurred in female rats at 75 mg/kg/day (days 23 to 30 postpartum) and in male rats at 200 mg/kg/day (day 37 to 44 postpartum) were not attributed to maternal treatment because the reductions were transient and independent of dose.
Sexual maturation:
no effects observed
Description (incidence and severity):
Sexual maturation in F1 rats was unaffected by treatment of P generation rats with the test substance at dosages as high as 500 mg/kg/day.
The average day on which vaginal opening was observed in F1 female rats was comparable among the dosage groups. In addition, the average body weight on the day sexual maturation was achieved in F1 male and female rats was comparable among the dosage groups and did not differ significantly from the vehicle control group values.
Organ weight findings including organ / body weight ratios:
effects observed, non-treatment-related
Description (incidence and severity):
There were no apparent effects of treatment of P generation rats on the reproductive and nonreproductive tissue weights of the F1 male or female rats at any dosage level tested.
A slight, but statistically significant increase (p≤0.01) in the absolute weight of the left epididymis occurred in F1 male rats at 500 mg/kg/day, in comparison to the vehicle control group value. This increase was considered unrelated to treatment of P generation rats because there was no statistically significant change in the contralateral organ or in the relative weight of the left epididymis. In addition, the statistically significant reduction (p≤0.01) in the relative (% terminal body weight) weight of the left kidney for F1 male rats at 75 mg/kg/day was not attributed to exposure to the test item because: 1) the reduction was independent of dose; and 2) there was no statistically significant change in the contralateral organ.
Gross pathological findings:
effects observed, non-treatment-related
Description (incidence and severity):
Preweaning:
There were no gross lesions observed in the F1 generation pups that survived to scheduled necropsy on day 22 postpartum. In the pups that were stillborn, found dead or humanely euthanized, no milk was present in the stomach of 5, 1, 1 and 1 F1 generation pups in the 0 (Vehicle), 75, 200 and 500 mg/kg/day dosage groups, respectively.
Postweaning:
There were no test substance-related necropsy observations. All necropsy observations were considered unrelated to treatment of P generation rats because: 1) the incidences were not dosage-dependent; and/or 2) the observations occurred in only one rat in any dosage group. In F1 male rats, these necropsy observations included numerous red areas on the thymus or left lateral lobe of the lung, a mottled (red and dark red) appearance to the right diaphragmatic lobe of the lung, an extra lobe present on the left lateral lobe of the liver, an enlarged right testis, slight dilation of the pelvis in the right kidney, a pale appearance to the left kidney and a clear fluid-filled cyst on the left kidney. Gross lesions observed in F1 female rats included numerous red areas on the thymus, a dark red area on the right diaphragmatic lobe of the lungs and slight dilation of the pelvis in the right kidney. No other gross lesions
occurred.
Other effects:
effects observed, non-treatment-related
Description (incidence and severity):
The lactation index was significantly increased (p≤0.05 or p≤0.01) in the 75 and 500 mg/kg/day dosage groups, in comparison to the vehicle control group value.
This increase was considered unrelated to treatment because: 1) the increase was independent ofdose; and/or 2) the value was within the historical range of the Testing Facility.
Anogenital distance on days 1 or 22 postpartum in F1 male and female pups was not affected by treatment of P generation rats with the test item at any dosage level tested.
Nipple eruption did not occur in any male pup at any dosage level tested. All female pups had nipples present on day 12 postpartum, with the exception of two from litter 16228 (F1 generation; 75 mg/kg/day dosage group).

Effect levels (F1)

Key result
Dose descriptor:
NOAEL
Remarks:
viability and growth
Generation:
F1
Effect level:
> 500 mg/kg bw/day
Based on:
test mat.
Sex:
male/female
Basis for effect level:
viability
body weight and weight gain

Overall reproductive toxicity

Key result
Reproductive effects observed:
no
Lowest effective dose / conc.:
500 mg/kg bw/day
Treatment related:
no

Applicant's summary and conclusion

Conclusions:
Based on the results of this study, the no-observable-adverse-effect-level (NOAEL) for toxicity of the test item is 200 mg/kg/day.
The reproductive NOAEL in the P generation rats and the NOAEL for viability and growth of the F1 generation offspring is greater than 500 mg/kg/day. There were no apparent effects on estrous cycling, mating and fertility, reproductive organ weights or natural delivery parameters in the P generation and growth and development (including anogenital distance, nipple eruption or sexual maturation) in the F1 generation rats at the highest dosage level tested (500 mg/kg/day).