Registration Dossier

Administrative data

Key value for chemical safety assessment

Effects on fertility

Description of key information
 An OECD 421 screening study for reproductive/development toxicity on the supporting substance dimethyl 1,4-cyclohexanedicarboxylate (DMCD) CAS No. 94-60-0 indicates the test material will have no adverse effects on reproductive or developmental parameters.
Link to relevant study records

Referenceopen allclose all

Endpoint:
screening for reproductive / developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
from September 10, 2002 to May 25, 2003
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 421 (Reproduction / Developmental Toxicity Screening Test)
Version / remarks:
Adopted July 27, 1995
Deviations:
no
Principles of method if other than guideline:
not applicable
GLP compliance:
yes
Limit test:
no
Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS:
-Strain: Crl: CD®(SD) IGS Br strain
-Source: Charles River Laboratories, Stone Ridge (Kingston), NY, USA
-Sex: male and female
-Age at study initiation: males - 64 days, females - 67 days
-Acclimation period: 5 days
-Weight at study receipt: males 317.64 ± 11.95 g; females 232.72 ± 10.44 g
-Housing: Animals were housed singly in suspended stainless steel wire mesh cages for the duration of the study except during the matling phase during which time animals were housed in pairs. Male and female cages were housed and racked separately except during mating. Soild bottom pans containing bedding material were placed in the cages of female rats on the 19th day of gestation. Cages and racks were washed approximately once a week. Absorbant paper used to collect excreta, was changed daily throughout the study for males and daily for female rats during the pre-mating, mating and gestation phases of the study. During the lactation phase of the study bedding material in the solid bottom pans was changed only if necessary.
Due to a detectable odour from the test substance each dose group was housed in a separate animal room to reduce the potential of airborne cross-contamination. No other studies were housed in these rooms during this study.
-Diet: Certified Rodent Diet (Purina Rodent Chow #5002, meal (PMI Feed, Inc. Richmond, IN USA) available ad libitum
-Water: local ad libitum
-Method of animal identification: unique eartag
-Method of animal distribution: Stratified randomisation program

ENVIRONMENTAL CONDITIONS:
-Temperature (°C): 20.9 -33.7
-Humidity (%): 29.3 - 67.9
-Photoperiod: 12 hours light/dark
There were several excursions above the recommended limit of 25°C, most of which were transient (90 minutes or less). However two excursions lasting approximately 24 hours occurred in the roommshousing the low and high dose groups (2002-10-02 to 2002-10-03). During this time the animals housed in these rooms were temporarily relocated to the room housing the mid-dose group and were removed back to their respective rooms once the temperature returned to an acceptable level.

IN-LIFE DATES:
-Study Initiation Date: September 10, 2002
-Experimental Start Date: September 16, 2002
-Experimental Completion Date: May 25, 2003
Route of administration:
oral: feed
Vehicle:
unchanged (no vehicle)
Details on exposure:
Test formulations:
The test substance was mixed with the ground food to test concentrations of 15.0, 4.5 and 1.5 mg test substance per gram of feed. The mixtures were prepared five or six times and used within 14 days based on the stability of the mixture. The control animals were given the basal diet.

Stability of the test substance in test diets was determined by repeated analysis of the high (15.0 mg/g) and low dose (1.5 mg/g) mixtures on days 0, 4, 7, 10, 14, 17 and 22 days. The 15 mg/g mixture was stable after 14 days but the 1.5 mg/g mixture was not.

Where the mean analytical concentration of the mixture was <10% of the target concentration new batches were prepared and the animals provided with new feeders.

Homegeneiy of the test mixtures was evauated by measuring the concentration of the test substance in Batch 1 at three levels (top, middle and bottom of the container) for all three dose levels. It was determined that at room temperature the cis isomer is a liquid but the trans isomer is a solid. Therefore container #2 of the test material was heated slightly to ensure the isomers were mixed and then this was used to prepare all the test mixtures for the studies. Purity of the test substance was analysed at the beginning and end of the study.

Exposure:
The study consisted of four phases: pre-mating (14 days), mating (1 to 14 days), pregnancy (21-23 days) and early lactation (4-6 days). Male rats were treated throughout the study (50 days). Female rats were treated throughout the study until they were euthanised (38 to 57 days). Males were euthanised on day 51. Female rats that delivered a litter and their offspring were euthanised on days 4, 5 or 6 postpartum. Female rats that showed evidence of mating but did not deliver were euthanised on day 23 of gestation.
Details on mating procedure:
Pairing was 1 male with 1 female within the same treatment group for 1 to 14 days. Mating was confirmed either from a sperm positive vaginal smear or a copulatory plug. The day of observation of a copulatory plug in situ and/or sperm in the lavage was designated as Gestation Day 0.

Following copulation male and female rats were separated and housed individually until study termination.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
The structure of the test material was confirmed using gas chromatography with mass spectrometric determination (GC/MS). Purity of the test substance was determined by gas chromatography with flame ionisation detection (GC/FID). Purity was confirmed at the start and end of the test period.

Stability of the test substance in test diets was determined by repeated analysis of the high (15.0 mg/g) and low dose (1.5 mg/g) mixtures on days 0, 4, 7, 10, 14, 17 (1.5 mg/g mixture only) and 22 days. The 15 mg/g mixture was stable after 14 days but the 1.5 mg/g mixture was not.

Where the mean analytical concentration of the mixture was <10% of the target concentration new batches were prepared and the animals provided with new feeders.

Homegeneiy of the test mixtures was evauated by measuring the concentration of the test substance in Batch 1 at three levels (top, middle and bottom of the container) for all three dose levels. It was determined that at room temperature the cis isomer is a liquid but the trans isomer is a solid. Therefore container #2 of the test material was heated slightly to ensure the isomers were mixed and then this was used to prepare all the test mixtures for the studies. Purity of the test substance was analysed at the beginning and end of the study.
Duration of treatment / exposure:
Females: 38 to 57 days
Males: 50 days
Frequency of treatment:
Daily
Details on study schedule:
In-Life Study Dates:
-Study Initiation Date: September 10, 2002
-Experimental Start Date: September 16, 2002
-Experimental Completion Date: May 25, 2003
Dose / conc.:
1 500 mg/kg diet
Remarks:
Dose level of approximately 92 mg/kg bw/day males and 111 mg/kg bw females
Dose / conc.:
4 500 mg/kg diet
Remarks:
Dose level of approximately 276 mg/kg bw/day males and 351 mg/kg bw females
Dose / conc.:
15 000 mg/kg diet
Remarks:
Dose level of approximately 888 mg/kg bw/day males and 1124 mg/kg bw females
No. of animals per sex per dose:
12 rats/sex/group
Control animals:
yes, plain diet
Details on study design:
Dose selection rationale:
Doses were selected in consultation with the sponsor based on the data from a previous repeated-dose oral toxicity study.

Doses received:
Males: 0, 92, 276 and 888 mg/kg bw/day
Females: 0, 111, 351 and 1124 mg/kg bw/day (time-weight average of dose levels for all phases of the study)
Positive control:
none
Parental animals: Observations and examinations:
Clinical Observations:
Clinical observations were conducted daily. Animals were also observed for moribundity/mortality each weekday afternoon. Clinical observations included but were not limited to: examination of hair, skin, eyes, mucous membranes, motor activity, feaces, urine, respiratory system, circulatory system, autonomic nervous system, central nervous system and behaviour patterns.

Body Weights:
Males: measured on days 0, 7 and weekly thereafter.
Females: measured on days 0, 7 and 14 of pre-mating phase, days 7 and 14 of mating (where applicable), days 0, 7, 14 and 20 of gestation, and days 0 and 4 postpartum.
Live pups: measured as a group on days 0 and 4 postpartum.

Food Consumption:
Feeders were weighed on days 0, 3, 7 and 14 of the premating phase for all animals. Feed consumption was not determined during mating. For male rats after mating the feeders were weighed once a week.. Additional feeder weights were collected for male rats from the mid- and high- dose groups on day 44 because a new batch of feed was prepared. For females feeders were weighed on days 0, 7, 14 and 20 of gestation, on days 0 and 4 postpartum and whenever feeders needed to be changed due to the stability of the test diet.

Water consumption:
Not monitored.
Oestrous cyclicity (parental animals):
not examined
Sperm parameters (parental animals):
Motility Evaluation
On the day of necropsy, percentage motility of sperm for each animal as taken from the right epididymis was determined.

Testicular Spermatid Head Counts
Homogenisation resistant spermatid head counts were performed on the frozen left testes. Total sperm counts were determined for each animal. Data were reported as total counts (million/organ) and adjusted for organ weight (thousand/mg tissue).

Epididymal Spermatozoan Counts
Homogenisation resistant spermatiozoan counts were performed on the frozen left epididymides (caudal left epididymis). Data were reported as total counts (million/organ) and adjusted for ogan weight (thousand/mg tissue).
Litter observations:
Clinical Observations:
All pups were observed once daily for clinical observations, moribundity and mortality. The number of live pups born and the number found dead in each litter were recorded. The live pups were sexed, counted and weighed.

Body Weights:
Body weights for live pups were measured as a group by sex on Postnatal Days 0 and 4.
Postmortem examinations (parental animals):
Adult males were anesthetised by carbon dioxide and euthanised by exsanguination . Adult females and pups were euthanised with carbon dioxide.
Adult males were fasted overnight prior to necropsy on day 51. Adult females that showed evidence of mating but no litter were necropsied on day 23 of gestation. All other female rats were necropsied on days 4, 5 or 6 postpartum.
Male rat terminal body weights were collected following exsangination. Terminal body weights were not taken for adult females.

All adult animals were subjected to necropsy and a gross examination with special attention to the reproductive organs.The uteri from all adult females were examined for implantation sites, Corpora lutea were also counted.
The following tissues from adult animals were fixed in 10% buffered formalin: ovaries, uterus, vagina, Fallopian tubes, male accessory sex glands and gross lesions. After mortality analysis the right testis and right epididymis were fixed in Bouin's fixative; after approximately 24 hours, these tissues were rinsed twice with 50% ethyl alcohol and then stored in 70% ethyl alcohol.
For long term storage, the right testis and right epididymis were stored in 10% buffered formalin. The left testis and left epididyis were placed into individual containers, frozen with dry ice and stored at -70°C.

Organ weights:
For males, the epididymides and testes were weighed; paired organs were weighed together.

Histopathology:
The overies, right testis and right epididymis were examined microscopically for animals from the control and high-dose groups. Collected gross lesions were also examined microscopically for animals from all dose groups. Tissues were sectioned with a rotary microtome set at a thickness of either 5 or 6 µm. the resulting tissue sections were stained with hematoxylin and oesin (H&E) and examined for histopathology. In addition, duplicate sections of testicular tissue were prepared and stained with the periodic acid-Schiff procedure (PAS) and examined for histopathology.
Postmortem examinations (offspring):
The pups were euthanised with carbon dioxide.
Statistics:
Adult body weight and body weight change, litter weight, pup body weight and weight change, feed consumption, feed utilisation, organ weights, organ-to-body weight ratios, precoital interval, gestation duration, numbers of implants and corpora lutea, pre- and post-implantation loss, live-birth index, numbers of live and dead pups, numbers of male and female pups, percent of male pups, and pup survival data were evaluated using Bartlett's test (p≤0.01), one-way analysis of variance (ANOVA) (p≤0.05).

Where variances of the means were not considered equal by the Bartlett's test (p≤0.01) the data were evaluated using a Kruskal-Wallis H-test (p≤0.05) followed by Mann-Whitney U-test (p≤0.05).

The reproductive performance, litter size, and the fertility and fecundity indices were evaluated in contingency tables using a Chi-square test (p≤0.05).

Reproductive indices:
Fertility Index = (Number pregant females * 100)/ Number cohabited pairs

Fecundity Index = (Number pregnant females * 100) / Number mated pairs with copulatory plug or sperm

Offspring viability indices:
Live Birth Index = (Number of pups born alive * 100) / Number of pups in the litter


Clinical signs:
effects observed, non-treatment-related
Description (incidence and severity):
Minimal reductions in the amount of faeces or softened faeces, which were observed for five rats in the high-dose group for one to two days of the study. Other abnormalities observed were considered to be incidental to exposure.
Mortality:
no mortality observed
Body weight and weight changes:
effects observed, non-treatment-related
Description (incidence and severity):
Mean feed consumption values and body weights for males and females in the high-dose groups were lower than the control group.
Food consumption and compound intake (if feeding study):
effects observed, non-treatment-related
Description (incidence and severity):
Mean feed consumption values and body weights for males and females in the high-dose groups were lower than the control group.
Food efficiency:
not specified
Water consumption and compound intake (if drinking water study):
not specified
Ophthalmological findings:
not examined
Haematological findings:
not examined
Clinical biochemistry findings:
not examined
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
no effects observed
Histopathological findings: non-neoplastic:
no effects observed
Histopathological findings: neoplastic:
no effects observed
Other effects:
no effects observed
Reproductive function: oestrous cycle:
not examined
Reproductive function: sperm measures:
no effects observed
Reproductive performance:
no effects observed
Clinical observations and mortality:
No mortality was observed during the study. There were no clinical observations that could be attributed to treatment with the test substance.

Body weights:
Lower (p≤0.05) mean body weights were observed for male rats from the high-dose group beginning on day 7 and continuing until study termination. Mean body weights also were lower (p≤0.05) for female rats from the high-dose group on day 20 of gestation when compared with the control group. Lower (p≤0.05) mean body weight gains were observed for male rats from all treated groups on day 7, and for male rats from the mid- and high-dose groups on day 42. mean body weight gains were also lower (p≤0.05) for female rats from the high-dose group on day 7 of the premating phase and on day 20 of gestation.

Although mean body weight gains were significantly higher (p≤0.05) for female rats from all treated groups on day 7 of gestation, this may be an artifact of lower control group values due to an inadvertant with-holding of feed for one day from the control group.

Overall mean bodyweight gains during gestation were comparable amongst groups.

Food consumption:
Lower (p≤0.05) mean food consumption values were observed for male rats from the high-dose group on days 7, 14, 35, 42, and 49 when compared with the control group. Mean feed consumption values were also lower (p≤0.05) for female rats from the high-dose group on days 7 of the pre-mating phase and day 14 of the gestation phase and for the mid-dose group on day 7 of gestation. Lower (p≤0.05) mean feed utilisation values were observed for male rats from the mid- and high-dose groups on days 7and 42. Mean feed utilisation values were also lower (p≤0.05) for female rats from the high-dose group on day 7 of the premating phase.
Although mean feed utilisation values were significantly higher (p≤0.05) for female rats from all treated groups on day 7 of gestation this may be an artifact of lower control group values due to an inadvertant with-holding of feed for one day from the control group.

Test substance intake:
No effects.

Reproductive performance:

Organ weights:
Mean terminal body weights were lower (p≤0.05) for male rats from the high-dose group. However, there were no differences in mean absolute and relative (to body weight) organ weights for any of the groups.

Gross pathology:
No treatment-realted gross leasions observed. All gross lesions observed considered incidental to exposure.

Histopathology:
No treatment-related changes.

Assessment of sperm morphology and development
No treatment-related effects.
Key result
Dose descriptor:
NOAEL
Effect level:
1 124 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: Highest dose tested in females was selected as pregnant females are considered to be more sensitive than males and as feed consumption patterns in males appears to be food refusal.
Key result
Critical effects observed:
no
Clinical signs:
no effects observed
Mortality / viability:
no mortality observed
Body weight and weight changes:
no effects observed
Food consumption and compound intake (if feeding study):
not examined
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
not examined
Clinical biochemistry findings:
not examined
Urinalysis findings:
not examined
Sexual maturation:
not examined
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
not examined
Histopathological findings:
not examined
Other effects:
no effects observed
Developmental immunotoxicity:
not examined
The duration of the gestation phase was shorter (p≤0.05) for female rats from the mid-dose group when compared with the control group. All other parameters were comparable amongst the groups.

Clinical signs:
No effects; abnormalities occurred with similar frequency in all treated and the control groups.

Body weight:
Mean pup weight change and percent pup weight change for days 0 to 4 were significantly (p≤0.05) higher for pups from the low-dose group when compared with the control group.
Key result
Dose descriptor:
NOAEL
Generation:
F1
Effect level:
1 124 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: no adverse effect at the highest dose tested
Key result
Critical effects observed:
no
Key result
Reproductive effects observed:
no

none

Conclusions:
The no-observed-effect-level for reproductive and neonatal toxicity was considered to be 1124 mg/kg bw/day (females), the highest concentration level tested.
Executive summary:

In a reproductive/developmental toxicity screening study performed according to the OECD test guideline No. 421 and in compliance with GLP, 12 rats/sex/group were exposed ad libitum in the diet to dimethyl 1,4-cyclohexanedicarboxylate at dose concentrations of 0, 0.15% ( approximately 92 mg/kg bw/day males and 111 mg/kg bw females), 0.45% (approximately 276 mg/kg bw/day males and 351 mg/kg bw females) and 1.5% (888 mg/kg bw/day males and 1124 mg/kg bw female)test substance in feed for 38 - 57days (females) or 50 days (males). 

There were no treatment-related effects on male or female fertility, pregnancy performance, or pup development in this reproductive and developmental toxicity screening study. There were no adverse effects on duration of gestation, gestation index, mean number of implant sites per pregnancy, mean number of pups born per litter, mean number of live pups per litter, live birth index, fertility and fecundity indexes, and group mean litter weights. No treatment-related abnormalities were observed in the pups. Although the duration of geatation phase was shorter for female rats in the mid-dose group there was no apparent effect on pup viability. The increased mean pup weight change and percentage pup weight change from days 0 to 4 of the low-dose group in comparison to the control group were not considered to be biologically significant. The no-observed-effect-level for reproductive and neonatal toxicity was considered to be 1124 mg/kg bw/day (females), the highest concentration level tested.

Under the conditions of the study, there was no clear evidence of test substance-related effects on reproduction of parental males and females or on survival and development of F1 pups. Based on this information, dimethyl 1,4-cyclohexanedicarboxylate is not a reproductive toxicant and is not selectively toxic to the developing fetus. Dimethyl 1,4-cyclohexanedicarboxylate is not classified for Developmental or Reproductive Toxicity according to the Annex VI of the Regulation (EC) No. 1272/2008 (CLP).

Endpoint:
screening for reproductive / developmental toxicity
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Justification for type of information:
REPORTING FORMAT FOR THE ANALOGUE APPROACH
See Iuclid section 13 for read-across justification

1. HYPOTHESIS FOR THE ANALOGUE APPROACH
This read-across is based on the hypothesis that source and target substances have similar toxicological properties because they are hydrolysed to common compounds by tissue esterases:
- The source 1 substance is predicted to be hydrolysed to Cyclohexane-1,4-dicarboxylic acid (1,4-CHDA = Source 2) and in ethanol,
- The target substance is predicted to be hydrolysed to Cyclohexane-1,4-dicarboxylic acid (1,4-CHDA = Source 2) and in methanol.

2. SOURCE AND TARGET CHEMICAL(S) (INCLUDING INFORMATION ON PURITY AND IMPURITIES)
The target and the source substances are reaction mass, composed of two diastereoisomers (trans- and cis-isomers).
The target and the source 2 substances are esters, whereas the source 1 substance is an acid.

3. ANALOGUE APPROACH JUSTIFICATION
The target and the source 2 substances showed similar toxicity profile in the acute toxicity study by oral route and are expected to follow the same bio-transformation pathway (hydrolysis to source 2 substance and alcohol). In addition, the source substance 2 is expected to be a worst-case compared to the target substance considering the respective physico-chemical properties.
The study design (OECD 421, GLP, Ref. 29) is adequate and reliable for the purpose of the prediction based on read-across. The test material used represents the source 2 substance as described in the hypothesis in terms of purity and impurities. The results of the studies are adequate for the purpose of classification and labelling.
Therefore, based on the considerations above, it can be concluded that the results of the acute oral toxicity studies conducted in both the rat and the mouse with the source substance are likely to predict the properties of the target substance and are considered as adequate to fulfil the information requirement of Annex VIII, 8.7.1.

4. DATA MATRIX
See Iuclid section 13.
Reason / purpose:
read-across source
Reason / purpose:
read-across: supporting information
Clinical signs:
effects observed, non-treatment-related
Description (incidence and severity):
Minimal reductions in the amount of faeces or softened faeces, which were observed for five rats in the high-dose group for one to two days of the study. Other abnormalities observed were considered to be incidental to exposure.
Mortality:
no mortality observed
Body weight and weight changes:
effects observed, non-treatment-related
Description (incidence and severity):
Mean feed consumption values and body weights for males and females in the high-dose groups were lower than the control group.
Food consumption and compound intake (if feeding study):
effects observed, non-treatment-related
Description (incidence and severity):
Mean feed consumption values and body weights for males and females in the high-dose groups were lower than the control group.
Food efficiency:
not specified
Water consumption and compound intake (if drinking water study):
not specified
Ophthalmological findings:
not examined
Haematological findings:
not examined
Clinical biochemistry findings:
not examined
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
no effects observed
Histopathological findings: non-neoplastic:
no effects observed
Histopathological findings: neoplastic:
no effects observed
Other effects:
no effects observed
Reproductive function: oestrous cycle:
not examined
Reproductive function: sperm measures:
no effects observed
Reproductive performance:
no effects observed
Clinical observations and mortality:
No mortality was observed during the study. There were no clinical observations that could be attributed to treatment with the test substance.

Body weights:
Lower (p≤0.05) mean body weights were observed for male rats from the high-dose group beginning on day 7 and continuing until study termination. Mean body weights also were lower (p≤0.05) for female rats from the high-dose group on day 20 of gestation when compared with the control group. Lower (p≤0.05) mean body weight gains were observed for male rats from all treated groups on day 7, and for male rats from the mid- and high-dose groups on day 42. mean body weight gains were also lower (p≤0.05) for female rats from the high-dose group on day 7 of the premating phase and on day 20 of gestation.

Although mean body weight gains were significantly higher (p≤0.05) for female rats from all treated groups on day 7 of gestation, this may be an artifact of lower control group values due to an inadvertant with-holding of feed for one day from the control group.

Overall mean bodyweight gains during gestation were comparable amongst groups.

Food consumption:
Lower (p≤0.05) mean food consumption values were observed for male rats from the high-dose group on days 7, 14, 35, 42, and 49 when compared with the control group. Mean feed consumption values were also lower (p≤0.05) for female rats from the high-dose group on days 7 of the pre-mating phase and day 14 of the gestation phase and for the mid-dose group on day 7 of gestation. Lower (p≤0.05) mean feed utilisation values were observed for male rats from the mid- and high-dose groups on days 7and 42. Mean feed utilisation values were also lower (p≤0.05) for female rats from the high-dose group on day 7 of the premating phase.
Although mean feed utilisation values were significantly higher (p≤0.05) for female rats from all treated groups on day 7 of gestation this may be an artifact of lower control group values due to an inadvertant with-holding of feed for one day from the control group.

Test substance intake:
No effects.

Reproductive performance:

Organ weights:
Mean terminal body weights were lower (p≤0.05) for male rats from the high-dose group. However, there were no differences in mean absolute and relative (to body weight) organ weights for any of the groups.

Gross pathology:
No treatment-realted gross leasions observed. All gross lesions observed considered incidental to exposure.

Histopathology:
No treatment-related changes.

Assessment of sperm morphology and development
No treatment-related effects.
Key result
Dose descriptor:
NOAEL
Effect level:
1 124 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: Highest dose tested in females was selected as pregnant females are considered to be more sensitive than males and as feed consumption patterns in males appears to be food refusal.
Key result
Critical effects observed:
no
Clinical signs:
no effects observed
Mortality / viability:
no mortality observed
Body weight and weight changes:
no effects observed
Food consumption and compound intake (if feeding study):
not examined
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
not examined
Clinical biochemistry findings:
not examined
Urinalysis findings:
not examined
Sexual maturation:
not examined
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
not examined
Histopathological findings:
not examined
Other effects:
no effects observed
Developmental immunotoxicity:
not examined
The duration of the gestation phase was shorter (p≤0.05) for female rats from the mid-dose group when compared with the control group. All other parameters were comparable amongst the groups.

Clinical signs:
No effects; abnormalities occurred with similar frequency in all treated and the control groups.

Body weight:
Mean pup weight change and percent pup weight change for days 0 to 4 were significantly (p≤0.05) higher for pups from the low-dose group when compared with the control group.
Key result
Dose descriptor:
NOAEL
Generation:
F1
Effect level:
1 124 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: no adverse effect at the highest dose tested
Key result
Critical effects observed:
no
Key result
Reproductive effects observed:
no

none

Conclusions:
The no-observed-effect-level for reproductive and neonatal toxicity was considered to be 1124 mg/kg bw/day (females), the highest concentration level tested for the source and the target substances.
Executive summary:

In a reproductive/developmental toxicity screening study performed according to the OECD test guideline No. 421 and in compliance with GLP, 12 rats/sex/group were exposed ad libitum in the diet to dimethyl 1,4-cyclohexanedicarboxylate at dose concentrations of 0, 0.15% ( approximately 92 mg/kg bw/day males and 111 mg/kg bw females), 0.45% (approximately 276 mg/kg bw/day males and 351 mg/kg bw females) and 1.5% (888 mg/kg bw/day males and 1124 mg/kg bw female)test substance in feed for 38 - 57days (females) or 50 days (males). 

There were no treatment-related effects on male or female fertility, pregnancy performance, or pup development in this reproductive and developmental toxicity screening study. There were no adverse effects on duration of gestation, gestation index, mean number of implant sites per pregnancy, mean number of pups born per litter, mean number of live pups per litter, live birth index, fertility and fecundity indexes, and group mean litter weights. No treatment-related abnormalities were observed in the pups. Although the duration of geatation phase was shorter for female rats in the mid-dose group there was no apparent effect on pup viability. The increased mean pup weight change and percentage pup weight change from days 0 to 4 of the low-dose group in comparison to the control group were not considered to be biologically significant. The no-observed-effect-level for reproductive and neonatal toxicity was considered to be 1124 mg/kg bw/day (females), the highest concentration level tested.

Under the conditions of the study, there was no clear evidence of test substance-related effects on reproduction of parental males and females or on survival and development of F1 pups. Based on this information, dimethyl 1,4-cyclohexanedicarboxylate is not a reproductive toxicant and is not selectively toxic to the developing fetus.

The source and the target substances are not classified for Developmental or Reproductive Toxicity according to the Annex VI of the Regulation (EC) No. 1272/2008 (CLP).

Endpoint:
extended one-generation reproductive toxicity - basic test design (Cohorts 1A, and 1B without extension)
Data waiving:
study scientifically not necessary / other information available
Justification for data waiving:
the extended one-generation reproductive toxicity study does not need to be conducted because there are no results from available repeated dose toxicity studies that indicate adverse effects on reproductive organs or tissues, or reveal other concerns in relation with reproductive toxicity
Justification for type of information:
JUSTIFICATION FOR DATA WAIVING
The available repeated dose toxicity studies (i.e. a 28-day on the substance itself, a 90- day study and an OECD 421 screening study on an analogue) do not indicate adverse effects on reproductive organs or tissues or reveal other concerns in relation with reproductive toxicity.
Reason / purpose:
data waiving: supporting information
Reason / purpose:
data waiving: supporting information
Reason / purpose:
data waiving: supporting information
Reproductive effects observed:
not specified
Effect on fertility: via oral route
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
1 124 mg/kg bw/day
Study duration:
subacute
Species:
rat
Quality of whole database:
The key study is GLP-compliant and of high quality (Klimisch score = 1).
Effect on fertility: via inhalation route
Endpoint conclusion:
no study available
Effect on fertility: via dermal route
Endpoint conclusion:
no study available
Additional information

There are no reproductive toxicity studies on the test material. A reproductive/development toxicity screening test performed according to OECD 421 is available on a supporting substance; Dimethyl 1,4-cyclohexanedicarboxylate (DMCD) CAS No. 94-60-0.  

Study summary:

12 rats/sex/group were exposed ad libitum in the diet to dimethyl 1,4-cyclohexanedicarboxylate at dose concentrations of 0, 0.15% ( approximately 92 mg/kg bw/day males and 111 mg/kg bw females), 0.45% (approximately 276 mg/kg bw/day males and 351 mg/kg bw females) and 1.5% (888 mg/kg bw/day males and 1124 mg/kg bw female) test substance in feed for 38 – 57 days (females) or 50 days (males). There were no treatment-related effects on male or female fertility, pregnancy performance, or pup development in this reproductive and developmental toxicity screening study. There were no adverse effects on reproductive or developmental parameters. Although the duration of the gestation phase was shorter for female rats in the mid-dose group there was no clear dose-response relationship. There was no apparent effect on pup viability. An increased mean pup weight change and percentage pup weight change from days 0 to 4 of the low-dose group in comparison to the control group was not considered to be biologically significant. The no-observed-effect-level for reproductive and neonatal toxicity was considered to be 1124 mg/kg bw/day (pregnant females more sensitive than males), the highest concentration level tested.   Under the conditions of the study, there was no clear evidence of test substance-related effects on reproduction of parental males and females or on survival and development of F1 pups.

Based on this information, the source and the target substances are not reproductive toxicant and are not selectively toxic to the developing foetus.

Effects on developmental toxicity

Description of key information

An OECD 414 pre-natal developmental toxicity study on the analogue substance dimethyl Cyclohexane-1,4-dicarboxylic acid, CAS No. 1076-97-7 indicates the test material will have no adverse effects on developmental parameters.

Link to relevant study records

Referenceopen allclose all

Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
30 January to 09 April 2015
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
Version / remarks:
Adopted 22 January 2001
Deviations:
no
Qualifier:
according to
Guideline:
EPA OPPTS 870.3700 (Prenatal Developmental Toxicity Study)
Principles of method if other than guideline:
Not applicable
GLP compliance:
yes
Limit test:
no
Species:
rat
Strain:
other: Sprague Dawley [Crl:CD(SD)]
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories, Inc., Raleigh, NC
- Age at receipt: Approximately 80 days
- Weight at study initiation: 214-276 g on gestation day 0
- Housing: All rats were housed 2-3 per cage in clean, solid-bottom cages with bedding material (Bed-O'Cobs®; The Andersons, Cob Products Division, Maumee, OH). Following positive evidence of mating, the females were individually housed in clean, solid-bottom cages with bedding material.
- Diet: Basal diet (PMI Nutrition International, LLC Certified Rodent LabDiet® 5002), ad libitum
- Water: Reverse osmosis-purified (on-site) drinking water, delivered by an automatic watering system, ad libitum
- Acclimation period: 12 days

ENVIRONMENTAL CONDITIONS
- Temperature: 70.0-70.8 °F (21.1-21.6 °C)
- Humidity: 39.3-50.5 %
- Air changes: A minimum of 10 fresh air changes per hour
- Photoperiod: Fluorescent lighting provided illumination for a 12 h light / 12 h dark photoperiod
Route of administration:
oral: gavage
Vehicle:
corn oil
Details on exposure:
PREPARATION OF DOSING SOLUTIONS:
- The test item formulations were prepared approximately weekly as single formulations for each dosage level, divided into aliquots for daily dispensation, and stored at room temperature, protected from light. The test item formulations were stirred continuously throughout the preparation, sampling, and dose administration procedures.
- The vehicle was dispensed approximately weekly for administration to the control group (Group 1) and for preparation of the test item formulations; aliquots were prepared for daily dispensation to the control group and stored at room temperature, protected from light. The vehicle was mixed throughout the sampling and dose administration procedures.

VEHICLE
- Concentration in vehicle: 50, 100 and 200 mg/mL
- Amount of vehicle (if gavage): 5 mL/kg bw/day
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Analyses to demonstrate the stability, homogeneity, and resuspension homogeneity of the test item formulations following 11 days of room temperature and refrigerated storage at target concentrations of 10 and 250 mg/mL were assessed in a previous study (Stinson, 2015, WIL-387057).
- Samples for homogeneity determinations were collected from the top, middle, and bottom strata of the first 50 and 200 mg/mL dosing formulations prepared at the smallest and largest formulation volumes prepared during the study. In addition, samples for resuspension homogeneity determinations were collected from the top and bottom strata of an aliquot taken from these same dosing formulations following refrigerated and room temperature storage for 11 days; aliquots were mixed for a minimum of 30 minutes prior to sample collection. Samples for concentration analysis were also collected from the middle stratum of the first and last dosing formulations at each dosage level (including the control group) prepared during the study. One set of samples from each collection was subjected to the appropriate analyses.
- All remaining samples were stored refrigerated (approximately 2-8 °C) as back-up. All analyses were conducted by the WIL Research Analytical Chemistry Department using a validated gas chromatography method using flame ionization detection (Stinson, 2015, WIL-387057).
Details on mating procedure:
- Impregnation procedure: Cohoused
- If cohoused:
- M/F ratio per cage: 1:1
- Selected females were approximately 13 weeks when paired for breeding
- Proof of pregnancy: Positive evidence of mating was confirmed by the presence of a vaginal copulatory plug or the presence of sperm in a vaginal lavage referred to as Day 0 of pregnancy.
Duration of treatment / exposure:
From gestation days 2 through 19
Frequency of treatment:
Once daily
Duration of test:
From gestation days 2 through 19.
Dose / conc.:
250 mg/kg bw/day (actual dose received)
Dose / conc.:
500 mg/kg bw/day (actual dose received)
Dose / conc.:
1 000 mg/kg bw/day (actual dose received)
No. of animals per sex per dose:
25 bred females/dose
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: Dosage levels were determined from results of a previous OECD 421 dietary study in rats. Dietary exposure to levels of DMCD of 0, 0.15, 0.45, or 1.5 % resulted in dosage levels of 0, 111, 351, or 1124 mg/kg bw/day for female rats over the course of the study. Reductions in feed consumption, body weights, and/or rate of body weight gain were noted in the female high-dosage group during the first 7 days of dietary consumption and feed consumption was decreased in the mid- and high-dosage groups on gestation day 7. Mean body weight and body weight gain was reduced in the high-dosage group on gestation day 20. No other effects were noted that were pertinent in setting dosage levels for the current OECD 414 study. It is not known if the reductions in the highest dosage group during the first few days of dosing were due to palatability issues with the treated feed. Based on these results, dosage levels were selected to test the limit dose and below as required by the testing guideline of this study.
- Rationale for animal assignment: The bred females were assigned to groups using a WTDMS™ computer program which randomized the animals based on stratification of the gestation day 0 body weights in a block design. Animals not assigned to study were transferred to the WIL Research stock colony or euthanized by carbon dioxide inhalation and discarded.
Maternal examinations:
CLINICAL OBSERVATIONS: Yes
- Time schedule: All rats were observed twice daily, once in the morning and once in the afternoon, for moribundity and mortality. Individual clinical observations were recorded daily from gestation days 0 through 20 (prior to dose administration during the treatment period). Animals were also observed for signs of toxicity approximately 1 hour following dose administration.

BODY WEIGHT: Yes
- Time schedule for examinations: Individual maternal body weights were recorded on gestation days 0 and 2-20 (daily). Group mean body weights were calculated for each of these days. Mean body weight changes were calculated for each corresponding interval and also for gestation days 2-6, 6-9, 9-12, 12-15, 15-20, and 2-20.

FOOD CONSUMPTION: Yes
- Individual food consumption was recorded on gestation days 0 and 2-20 (daily). Food intake was reported as g/animal/day and g/kg/day for the corresponding body weight change intervals. When food consumption could not be determined for an animal during a given interval (due to a weighing error, food spillage, etc.), group mean values were calculated for that interval using the available data.

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day 20; All females were euthanized on gestation day 20 by carbon dioxide inhalation.
- The cranial, thoracic, abdominal, and pelvic cavities were opened by a ventral mid-line incision, and the contents were examined. In all instances, the postmortem findings were correlated with the antemortem observations, and any abnormalities were recorded.
- Organs examined: Following organ weight collection, the brain, liver, spleen, and thymus gland were preserved in 10 % neutral-buffered formalin for possible future histopathologic examination; other maternal tissues were preserved only as indicated by the gross findings. Representative sections of corresponding organs from a sufficient number of control animals were retained for comparison. The carcass of each female was then discarded.

OTHER:
- Gravid uterine weight was collected and net body weight (the gestation day 20 body weight exclusive of the weight of the uterus and contents) and net body weight change (the gestation day 0-20 body weight change exclusive of the weight of the uterus and contents) were calculated and presented for each gravid female at the scheduled laparohysterectomy.
- Organ weights: The following organs were weighed from all maternal animals at the scheduled necropsy: brain, liver, spleen, and thymus gland were weighed. Absolute weights and organ to brain weight ratios were reported.
Ovaries and uterine content:
The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: Yes
- Number of corpora lutea: Yes
- Number of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes
- Other: Uteri with no macroscopic evidence of implantation were opened and subsequently placed in 10 % ammonium sulfide solution for detection of early implantation loss (Salewski, 1964).
Fetal examinations:
- Each viable fetus was examined externally, individually sexed, weighed, euthanized by a subcutaneous injection of sodium pentobarbital in the scapular region.
- External examinations: Yes; detailed external examination of each fetus included, but was not limited to, an examination of the eyes, palate, and external orifices, and each finding was recorded.
- Soft tissue examinations: Yes; Each viable fetus was subjected to a visceral examination using a modification of the Stuckhardt and Poppe fresh dissection technique to include the heart and major blood vessels (Stuckhardt and Poppe, 1984). The sex of each fetus was confirmed by internal examination. Fetal kidneys were examined and graded for renal papillae development (Woo and Hoar, 1972).
- Skeletal examinations: Yes
- Head examinations: Yes: Heads from approximately one-half of the fetuses in each litter were placed in Harrison’s fixative for subsequent soft-tissue examination by the Wilson sectioning technique (Wilson, 1965). The heads from the remaining one-half of the fetuses were examined by a midcoronal slice.
- All carcasses were eviscerated and fixed in 100% ethyl alcohol.
Following fixation in alcohol, each fetus was stained with Alizarin Red S (Dawson, 1926) and Alcian Blue (Inouye, 1976). Fetuses were then examined for skeletal malformations and developmental variations.
Statistics:
See "Any other information on materials and methods incl. tables"
Indices:
Group Mean Litter Basis:
Postimplantation Loss/Litter = [No. Dead Fetuses, Resorptions (Early/Late)/Group] / [No. Gravid Females/Group]

Proportional Litter Basis:
Summation Per Group (%) = [Sum of Postimplantation Loss/Litter (%)] / [No. Litters/Group]

Where:
Postimplantation Loss/Litter (%) = ([No. Dead Fetuses, Resorptions (Early/Late)/Litter] / [No. Implantation Sites/Litter]) x 100

Summation per Group (%) = [Sum of Viable Fetuses Affected/Litter (%)] / [No. Litters/Group]
Where:
Viable Fetuses Affected/Litter (%) = [No. Viable Fetuses Affected/Litter] / [No. Viable Fetuses/Litter] x 100
Historical control data:
Results were compared with historical control data of the testing laboratory.
Clinical signs:
effects observed, non-treatment-related
Mortality:
no mortality observed
Body weight and weight changes:
effects observed, non-treatment-related
Food consumption and compound intake (if feeding study):
effects observed, non-treatment-related
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
not examined
Clinical biochemistry findings:
not examined
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
effects observed, non-treatment-related
Gross pathological findings:
no effects observed
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
no effects observed
Histopathological findings: neoplastic:
no effects observed
Other effects:
no effects observed
Number of abortions:
no effects observed
Pre- and post-implantation loss:
no effects observed
Total litter losses by resorption:
no effects observed
Early or late resorptions:
no effects observed
Dead fetuses:
no effects observed
Changes in pregnancy duration:
no effects observed
Description (incidence and severity):
Migrated Data from removed field(s)
Field "Effects on pregnancy duration" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsMaternalAnimals.MaternalDevelopmentalToxicity.EffectsOnPregnancyDuration): no effects observed
Changes in number of pregnant:
no effects observed
Other effects:
no effects observed
Details on maternal toxic effects:
Maternal toxic effects:no effects

Details on maternal toxic effects:
MATERNAL CLINICAL OBSERVATIONS AND SURVIVAL
- All females in the control, 250, 500, and 1000 mg/kg bw/day groups survived to the scheduled necropsy on gestation day 20. Test item-related increased incidences of clear and red material around the mouth were noted for all test item-treated groups approximately 1 hour following dose administration generally throughout the treatment period (gestation days 2-20). These clinical findings did not persist to the daily examinations and were therefore not considered adverse. No test item-related clinical findings were noted at the daily examinations at any dosage level. Findings noted in the test item-treated groups, including hair loss on the forelimbs and red material around the nose, occurred infrequently, at similar frequencies in the control group, and/or in a manner that was not dose-related.

MATERNAL BODY WEIGHTS AND GRAVID UTERINE WEIGHTS
- Slightly lower mean body weight gains were noted for all test item-treated groups compared to the control group during gestation days 2 through 6; the differences in the 250 and 1000 mg/kg bw/day groups were significant (p<0.05 or p<0.01) for the gestation day 2-6 interval, but did not occur in a dose-responsive manner. Mean body weight gains in the test item-treated groups were similar to the control throughout the remainder of the treatment period (gestation days 6-20) and when the overall treatment period (gestation days 2-20) was evaluated. In addition, mean body weights in the test item-treated groups were similar to the control group throughout the study. Therefore, the initial lower mean body weight gains observed in the 250, 500, and 1000 mg/kg bw/day groups were considered test item-related; however, because they were transient in nature they were not considered adverse. A significantly (p<0.01) lower mean net body weight gain was noted for the 1000 mg/kg bw/day group. Because there were no effects on mean body weights or mean net body weight in this group, the lower mean net body weight gain in the 1000 mg/kg bw/day group was not attributed to the test item. Mean net body weights and gravid uterine weights in the 250, 500, and 1000 mg/kg bw/day groups and net body weight gains in the 250 and 500 mg/kg bw/day groups were similar to the control group.

MATERNAL FOOD CONSUMPTION
- Maternal food consumption, evaluated as g/animal/day and g/kg/day, in the 250, 500, and 1000 mg/kg bw/day groups was unaffected by test item administration. The only significant (p<0.05 or p<0.01) differences from the control group were lower mean food consumption in the 500 and 1000 mg/kg bw/day groups during gestation day 7-8; these differences were transient and not considered to be test item-related.

MATERNAL NECROPSY DATA
- At the scheduled necropsy on gestation day 20, no test item-related internal findings were observed at dosage levels of 250, 500, and 1000 mg/kg bw/day. Macroscopic findings observed in the test item-treated groups occurred infrequently, at similar frequencies in the control group, and/or in a manner that was not dose-related. Five, 5, 5, and 3 females in the control, 250, 500, and 1000 mg/kg bw/day groups, respectively, were determined to be nongravid; 3, 2, 2, and 3 of these females, respectively, mated on the first day of pairing. Progressively fewer nongravid females were noted with each subsequent day of mating. The higher incidence of nongravid females was not attributed to the test item as mating occurred prior to test item administration; in addition, it occurred with equal representation in the control group.

ORGAN WEIGHTS
- Slightly lower (not statistically significant) mean absolute thymus gland weights were noted in all test item-treated groups. Due to the lack of statistical significance when compared to the control group and the absence of an effect on the mean thymus gland weight relative to brain weight; the differences in mean absolute thymus gland weights were not considered to be test item-related. Mean absolute and/or relative (to brain weight) brain, liver, and spleen weights in the 250, 500, and 1000 mg/kg bw/day groups were similar to the control group.

GESTATION DAY 20 LAPAROHYSTERECTOMY DATA
- Intrauterine growth and survival were unaffected by test item administration at dosage levels of 250, 500, and 1000 mg/kg bw/day. Parameters evaluated included postimplantation loss, live litter size, mean fetal body weights, and fetal sex ratios. Mean numbers of corpora lutea and implantation sites and the mean litter proportions of pre-implantation loss were similar across all groups. Differences from the control group were slight and not statistically significant.
Key result
Dose descriptor:
NOAEL
Effect level:
1 000 mg/kg bw/day (actual dose received)
Based on:
test mat.
Basis for effect level:
other: no adverse effect at the highest dose tested
Key result
Abnormalities:
no effects observed
Fetal body weight changes:
no effects observed
Description (incidence and severity):
Migrated Data from removed field(s)
Field "Fetal/pup body weight changes" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsFetuses.FetalPupBodyWeightChanges): no effects observed
Reduction in number of live offspring:
no effects observed
Changes in sex ratio:
no effects observed
Changes in litter size and weights:
no effects observed
Changes in postnatal survival:
no effects observed
External malformations:
effects observed, non-treatment-related
Skeletal malformations:
effects observed, non-treatment-related
Visceral malformations:
no effects observed
Other effects:
not specified
Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects:no effects

Details on embryotoxic / teratogenic effects:
FETAL MORPHOLOGICAL DATA
- The numbers of fetuses (litters) available for morphological evaluation were 247(20), 255(20), 270(20), and 283(22) in the control, 250, 500, and 1000 mg/kg bw/day groups, respectively. Malformations were observed in 0(0), 4(2), 2(2), and 0(0) fetuses (litters) in these same respective dose groups and were considered spontaneous in origin.

EXTERNAL MALFORMATIONS AND VARIATIONS
- Fetus no. 2343-09 in the 250 mg/kg bw/day group that had a vertebral agenesis (with a filamentous tail). Skeletally, all vertebrae posterior to sacral vertebra no. 1 were absent in this fetus. This finding was limited to a single fetus, not observed in a dose-related manner, and the mean litter proportion of the finding was within the range of values in the WIL Research historical control data and was not statistically significantly different from the concurrent control group. Therefore, vertebral agenesis in the 250 mg/kg bw/day group was not attributed to maternal test item administration.
- No other external malformations were noted for fetuses on this study. No external developmental variations were noted for any fetuses in this study.

VISCERAL MALFORMATIONS AND VARIATIONS
- No visceral malformations were noted for fetuses in this study. Visceral developmental variations noted in all group, including the control group, consisted of distended ureters and accessory lobule(s) in the liver. Other findings observed in the test item-treated groups, were noted infrequently and in a manner that was not dose-related. In addition, the mean litter proportions of these findings were not statistically significantly different from the control group and were within the ranges of the WIL Research historical control data. Therefore, the visceral developmental variations noted in 250, 500, and 1000 mg/kg bw/day groups were not considered test item-related.
- Dark red areas on the adrenal glands were noted for fetus nos. 2307-01 and 2307-11 in the control group and fetus no. 2371-03 in the 250 mg/kg bw/day group. In addition, dark red discoloration of the thyroid gland was noted for fetus no. 2307-01 in the control group and renal papilla(e) not fully developed (Woo and Hoar Grade 1) was noted for fetus nos. 2389-03 and 2389-08 in the 250 mg/kg bw/day group. These findings were not classified as either a malformation or developmental variation, were not included on the summary tables, and were not considered to be test item-related because they occurred infrequently, at similar frequencies in the control group, and/or in a manner that was not dose-related.

SKELETAL MALFORMATIONS AND VARIATIONS
- Skeletal malformations were noted for 0(0), 3(1), 2(2) and 0(0) fetuses (litters) in the control, 250, 500, and 1000 mg/kg bw/day groups, respectively. Rib anomalies were noted for fetus nos. 2389-05, 2389-06, and 2389-08 in the 250 mg/kg bw/day group. This finding consisted of fused ribs. In addition, fetus nos. 2389-05 and 2363-04 in the 250 and 500 mg/kg bw/day groups, respectively, had costal cartilage anomalies that consisted of misshapen or malpositioned costal cartilage. Fetus no. 2378-05 in the 500 mg/kg bw/day group had severely malaligned sternebrae. Skeletal malformations observed in the test item-treated groups occurred in single fetuses or litters, were not observed in a dose-related manner, and/or the mean litter proportions of the findings were within the WIL Research historical control value ranges and not statistically significantly different from the concurrent control group. Therefore, these findings were not attributed to maternal test item administration. No other skeletal malformations were noted at any dosage level.
- Skeletal developmental variations observed all groups, including the control group, consisted of 14th rudimentary rib(s), ossified cervical centrum no. 1, unossified sternebra(e) nos. 5 and/or 6, 7th cervical rib(s), reduced ossification of the 13th rib(s), unossified sternebra(e) nos. 1, 2, 3, and/or 4, and slightly or moderately malaligned sternebra(e). These findings were noted similarly in the control group and/or in a manner that was not dose-related. In addition, the mean litter proportions of these findings were not statistically significantly different from the concurrent control group and were within the WIL Research historical control data ranges. Other skeletal developmental variations were noted in single fetuses or litters, did not occur in a dose-related manner, and/or were within the WIL Research historical control data ranges. No relationship to the test item was evident for any skeletal developmental variation.
Key result
Dose descriptor:
NOAEL
Effect level:
1 000 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: no adverse effect at the highest dose tested
Key result
Abnormalities:
no effects observed
Key result
Developmental effects observed:
no

ANALYSES OF DOSING FORMULATIONS

The analyzed dosing formulations were within WIL Research’s SOP range for suspensions (85 to 115 %) and were homogeneous. The test item was not detected in the vehicle formulation that was administered to the control group (Group 1).

 

Table 7.8.2/1: Results of Homogeneity Analyses

Homogeneity Assessment

Low Group

(50 mg/mL)

High Group

(200 mg/mL)

Homogeneity Assessment of the 12-Feb-2015 Formulations

Mean Concentration (mg/mL)

50.5

196

RSD (%)

0.61

0.57

Mean % of Target

101

98.2

11-Day Refrigerated Storage Resuspension Homogeneity Assessment of the 12-Feb-2015 Formulations

Mean Concentration (mg/mL)

51.3

201

RSD (%)

1.1

1.0

Mean % of Target

103

100

11-Day Room Temperature Storage Resuspension Homogeneity Assessment of the 12-Feb-2015 Formulations

Mean Concentration (mg/mL)

51.0

199

RSD (%)

0.37

0.56

Mean % of Target

102

99.7

 

Table 7.8.2/2: Results of Concentration Analyses

Date of Preparation

Mean Concentration, mg/mL (% of Target)

Group 2

(50 mg/mL)

Group 3

(100 mg/mL)

Group 4

(200 mg/mL)

12-Feb-2015

50.6 (101)

99.9 (99.9)

197 (98.6)

27-Feb-2015

51.3 (103)

104 (104)

197 (98.6)

Conclusions:
Under the test conditions, the NOAEL for maternal toxicity and embryo/fetal development is 1000 mg/kg bw/day in rats.
Executive summary:

In a prenatal developmental toxicity study performed according to OECD Guideline 414 and in compliance with GLP, test substance was administered by oral (gavage) to groups of mated female Crl:CD(SD) rats (25/dose) at the dose levels of 0 (corn oil),250, 500, and 1000mg/kg bw/day from gestation days 2 through 19. The females were approximately 13 weeks of age at the initiation of dose administration. All animals were observed twice daily for mortality and moribundity. Clinical observations, body weights, and food consumption were recorded at appropriate intervals. On gestation day 20, a laparohysterectomy was performed on each female. The uteri, placentae, and ovaries were examined, and the numbers of fetuses, early and late resorptions, total implantations, and corpora lutea were recorded. Gravid uterine weights were recorded, and net body weights and net body weight changes were calculated. Selected organs were weighed. The fetuses were weighed, sexed, and examined for external, visceral, and skeletal malformations and developmental variations.

 

All females in the control, 250, 500, and 1000 mg/kg bw/day groups survived to the scheduled necropsy on gestation day 20. Test item-related clinical findings were limited to dose-dependent clear and red material around the mouth in all test item-treated groups approximately 1 hour following dose administration generally throughout the treatment period. These findings did not persist to the daily examinations (prior to dose administration) on the following day and were not considered to be adverse.

 

Test item-related slightly lower mean body weight gains were noted in the 250, 500, and 1000 mg/kg bw/day groups during gestation days 2-6. Mean body weight gains in the test item-treated groups were similar to the control group throughout the remainder of the treatment period (gestation days 6-19) and there were no corresponding effects on mean food consumption or mean absolute body weights. Therefore, the initial body weight decrements in the 250, 500, and 1000 mg/kg bw/day groups were not considered to be adverse. Mean maternal body weights, net body weights, net body weight gains, gravid uterine weights, and food consumption in the 250, 500, and 1000 mg/kg bw/day groups were unaffected by test item administration. No test item-related macroscopic findings or effects on brain, liver, thymus, and spleen weights were noted at any dosage level. Intrauterine growth and survival and fetal morphology in the 250, 500, and 1000 mg/kg bw/day groups were unaffected by maternal test item administration.

 

Based on the lack of adverse maternal or developmental toxicity observed at any dosage level, the NOAEL for maternal toxicity and embryo/fetal development is 1000 mg/kg bw/day in rats.
Endpoint:
developmental toxicity
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Justification for type of information:
REPORTING FORMAT FOR THE ANALOGUE APPROACH
See Iuclid section 13 for read-across justification

1. HYPOTHESIS FOR THE ANALOGUE APPROACH
This read-across is based on the hypothesis that source and target substances have similar toxicological properties because they are hydrolysed to common compounds by tissue esterases:
- The source 1 substance is predicted to be hydrolysed to Cyclohexane-1,4-dicarboxylic acid (1,4-CHDA = Source 2) and in ethanol,
- The target substance is predicted to be hydrolysed to Cyclohexane-1,4-dicarboxylic acid (1,4-CHDA = Source 2) and in methanol.

2. SOURCE AND TARGET CHEMICAL(S) (INCLUDING INFORMATION ON PURITY AND IMPURITIES)
The target and the source substances are reaction mass, composed of two diastereoisomers (trans- and cis-isomers).
The target and the source 2 substances are esters, whereas the source 1 substance is an acid.

3. ANALOGUE APPROACH JUSTIFICATION
The target and the source 2 substances showed similar toxicity profile in the acute toxicity study by oral route and are expected to follow the same bio-transformation pathway (hydrolysis to source 2 substance and alcohol). In addition, the source substance 2 is expected to be a worst-case compared to the target substance considering the respective physico-chemical properties.
The study design (OECD 414, GLP, Ref. 30) is adequate and reliable for the purpose of the prediction based on read-across. The test material used represents the source 2 substance as described in the hypothesis in terms of purity and impurities. The results of the studies are adequate for the purpose of classification and labelling.
Therefore, based on the considerations above, it can be concluded that the results of the acute oral toxicity studies conducted in both the rat and the mouse with the source substance are likely to predict the properties of the target substance and are considered as adequate to fulfil the information requirement of Annex IX, 8.7.2.

4. DATA MATRIX
See Iuclid section 13.
Reason / purpose:
read-across source
Reason / purpose:
read-across: supporting information
Clinical signs:
effects observed, non-treatment-related
Mortality:
no mortality observed
Body weight and weight changes:
effects observed, non-treatment-related
Food consumption and compound intake (if feeding study):
effects observed, non-treatment-related
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
not examined
Clinical biochemistry findings:
not examined
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
effects observed, non-treatment-related
Gross pathological findings:
no effects observed
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
no effects observed
Histopathological findings: neoplastic:
no effects observed
Other effects:
no effects observed
Number of abortions:
no effects observed
Pre- and post-implantation loss:
no effects observed
Total litter losses by resorption:
no effects observed
Early or late resorptions:
no effects observed
Dead fetuses:
no effects observed
Changes in pregnancy duration:
no effects observed
Description (incidence and severity):
Migrated Data from removed field(s)
Field "Effects on pregnancy duration" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsMaternalAnimals.MaternalDevelopmentalToxicity.EffectsOnPregnancyDuration): no effects observed
Changes in number of pregnant:
no effects observed
Other effects:
no effects observed
Details on maternal toxic effects:
Maternal toxic effects:no effects

Details on maternal toxic effects:
MATERNAL CLINICAL OBSERVATIONS AND SURVIVAL
- All females in the control, 250, 500, and 1000 mg/kg bw/day groups survived to the scheduled necropsy on gestation day 20. Test item-related increased incidences of clear and red material around the mouth were noted for all test item-treated groups approximately 1 hour following dose administration generally throughout the treatment period (gestation days 2-20). These clinical findings did not persist to the daily examinations and were therefore not considered adverse. No test item-related clinical findings were noted at the daily examinations at any dosage level. Findings noted in the test item-treated groups, including hair loss on the forelimbs and red material around the nose, occurred infrequently, at similar frequencies in the control group, and/or in a manner that was not dose-related.

MATERNAL BODY WEIGHTS AND GRAVID UTERINE WEIGHTS
- Slightly lower mean body weight gains were noted for all test item-treated groups compared to the control group during gestation days 2 through 6; the differences in the 250 and 1000 mg/kg bw/day groups were significant (p<0.05 or p<0.01) for the gestation day 2-6 interval, but did not occur in a dose-responsive manner. Mean body weight gains in the test item-treated groups were similar to the control throughout the remainder of the treatment period (gestation days 6-20) and when the overall treatment period (gestation days 2-20) was evaluated. In addition, mean body weights in the test item-treated groups were similar to the control group throughout the study. Therefore, the initial lower mean body weight gains observed in the 250, 500, and 1000 mg/kg bw/day groups were considered test item-related; however, because they were transient in nature they were not considered adverse. A significantly (p<0.01) lower mean net body weight gain was noted for the 1000 mg/kg bw/day group. Because there were no effects on mean body weights or mean net body weight in this group, the lower mean net body weight gain in the 1000 mg/kg bw/day group was not attributed to the test item. Mean net body weights and gravid uterine weights in the 250, 500, and 1000 mg/kg bw/day groups and net body weight gains in the 250 and 500 mg/kg bw/day groups were similar to the control group.

MATERNAL FOOD CONSUMPTION
- Maternal food consumption, evaluated as g/animal/day and g/kg/day, in the 250, 500, and 1000 mg/kg bw/day groups was unaffected by test item administration. The only significant (p<0.05 or p<0.01) differences from the control group were lower mean food consumption in the 500 and 1000 mg/kg bw/day groups during gestation day 7-8; these differences were transient and not considered to be test item-related.

MATERNAL NECROPSY DATA
- At the scheduled necropsy on gestation day 20, no test item-related internal findings were observed at dosage levels of 250, 500, and 1000 mg/kg bw/day. Macroscopic findings observed in the test item-treated groups occurred infrequently, at similar frequencies in the control group, and/or in a manner that was not dose-related. Five, 5, 5, and 3 females in the control, 250, 500, and 1000 mg/kg bw/day groups, respectively, were determined to be nongravid; 3, 2, 2, and 3 of these females, respectively, mated on the first day of pairing. Progressively fewer nongravid females were noted with each subsequent day of mating. The higher incidence of nongravid females was not attributed to the test item as mating occurred prior to test item administration; in addition, it occurred with equal representation in the control group.

ORGAN WEIGHTS
- Slightly lower (not statistically significant) mean absolute thymus gland weights were noted in all test item-treated groups. Due to the lack of statistical significance when compared to the control group and the absence of an effect on the mean thymus gland weight relative to brain weight; the differences in mean absolute thymus gland weights were not considered to be test item-related. Mean absolute and/or relative (to brain weight) brain, liver, and spleen weights in the 250, 500, and 1000 mg/kg bw/day groups were similar to the control group.

GESTATION DAY 20 LAPAROHYSTERECTOMY DATA
- Intrauterine growth and survival were unaffected by test item administration at dosage levels of 250, 500, and 1000 mg/kg bw/day. Parameters evaluated included postimplantation loss, live litter size, mean fetal body weights, and fetal sex ratios. Mean numbers of corpora lutea and implantation sites and the mean litter proportions of pre-implantation loss were similar across all groups. Differences from the control group were slight and not statistically significant.
Key result
Dose descriptor:
NOAEL
Effect level:
1 000 mg/kg bw/day (actual dose received)
Based on:
test mat.
Basis for effect level:
other: no adverse effect at the highest dose tested
Key result
Abnormalities:
no effects observed
Fetal body weight changes:
no effects observed
Description (incidence and severity):
Migrated Data from removed field(s)
Field "Fetal/pup body weight changes" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsFetuses.FetalPupBodyWeightChanges): no effects observed
Reduction in number of live offspring:
no effects observed
Changes in sex ratio:
no effects observed
Changes in litter size and weights:
no effects observed
Changes in postnatal survival:
no effects observed
External malformations:
effects observed, non-treatment-related
Skeletal malformations:
effects observed, non-treatment-related
Visceral malformations:
no effects observed
Other effects:
not specified
Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects:no effects

Details on embryotoxic / teratogenic effects:
FETAL MORPHOLOGICAL DATA
- The numbers of fetuses (litters) available for morphological evaluation were 247(20), 255(20), 270(20), and 283(22) in the control, 250, 500, and 1000 mg/kg bw/day groups, respectively. Malformations were observed in 0(0), 4(2), 2(2), and 0(0) fetuses (litters) in these same respective dose groups and were considered spontaneous in origin.

EXTERNAL MALFORMATIONS AND VARIATIONS
- Fetus no. 2343-09 in the 250 mg/kg bw/day group that had a vertebral agenesis (with a filamentous tail). Skeletally, all vertebrae posterior to sacral vertebra no. 1 were absent in this fetus. This finding was limited to a single fetus, not observed in a dose-related manner, and the mean litter proportion of the finding was within the range of values in the WIL Research historical control data and was not statistically significantly different from the concurrent control group. Therefore, vertebral agenesis in the 250 mg/kg bw/day group was not attributed to maternal test item administration.
- No other external malformations were noted for fetuses on this study. No external developmental variations were noted for any fetuses in this study.

VISCERAL MALFORMATIONS AND VARIATIONS
- No visceral malformations were noted for fetuses in this study. Visceral developmental variations noted in all group, including the control group, consisted of distended ureters and accessory lobule(s) in the liver. Other findings observed in the test item-treated groups, were noted infrequently and in a manner that was not dose-related. In addition, the mean litter proportions of these findings were not statistically significantly different from the control group and were within the ranges of the WIL Research historical control data. Therefore, the visceral developmental variations noted in 250, 500, and 1000 mg/kg bw/day groups were not considered test item-related.
- Dark red areas on the adrenal glands were noted for fetus nos. 2307-01 and 2307-11 in the control group and fetus no. 2371-03 in the 250 mg/kg bw/day group. In addition, dark red discoloration of the thyroid gland was noted for fetus no. 2307-01 in the control group and renal papilla(e) not fully developed (Woo and Hoar Grade 1) was noted for fetus nos. 2389-03 and 2389-08 in the 250 mg/kg bw/day group. These findings were not classified as either a malformation or developmental variation, were not included on the summary tables, and were not considered to be test item-related because they occurred infrequently, at similar frequencies in the control group, and/or in a manner that was not dose-related.

SKELETAL MALFORMATIONS AND VARIATIONS
- Skeletal malformations were noted for 0(0), 3(1), 2(2) and 0(0) fetuses (litters) in the control, 250, 500, and 1000 mg/kg bw/day groups, respectively. Rib anomalies were noted for fetus nos. 2389-05, 2389-06, and 2389-08 in the 250 mg/kg bw/day group. This finding consisted of fused ribs. In addition, fetus nos. 2389-05 and 2363-04 in the 250 and 500 mg/kg bw/day groups, respectively, had costal cartilage anomalies that consisted of misshapen or malpositioned costal cartilage. Fetus no. 2378-05 in the 500 mg/kg bw/day group had severely malaligned sternebrae. Skeletal malformations observed in the test item-treated groups occurred in single fetuses or litters, were not observed in a dose-related manner, and/or the mean litter proportions of the findings were within the WIL Research historical control value ranges and not statistically significantly different from the concurrent control group. Therefore, these findings were not attributed to maternal test item administration. No other skeletal malformations were noted at any dosage level.
- Skeletal developmental variations observed all groups, including the control group, consisted of 14th rudimentary rib(s), ossified cervical centrum no. 1, unossified sternebra(e) nos. 5 and/or 6, 7th cervical rib(s), reduced ossification of the 13th rib(s), unossified sternebra(e) nos. 1, 2, 3, and/or 4, and slightly or moderately malaligned sternebra(e). These findings were noted similarly in the control group and/or in a manner that was not dose-related. In addition, the mean litter proportions of these findings were not statistically significantly different from the concurrent control group and were within the WIL Research historical control data ranges. Other skeletal developmental variations were noted in single fetuses or litters, did not occur in a dose-related manner, and/or were within the WIL Research historical control data ranges. No relationship to the test item was evident for any skeletal developmental variation.
Key result
Dose descriptor:
NOAEL
Effect level:
1 000 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: no adverse effect at the highest dose tested
Key result
Abnormalities:
no effects observed
Key result
Developmental effects observed:
no

ANALYSES OF DOSING FORMULATIONS

The analyzed dosing formulations were within WIL Research’s SOP range for suspensions (85 to 115 %) and were homogeneous. The test item was not detected in the vehicle formulation that was administered to the control group (Group 1).

 

Table 7.8.2/1: Results of Homogeneity Analyses

Homogeneity Assessment

Low Group

(50 mg/mL)

High Group

(200 mg/mL)

Homogeneity Assessment of the 12-Feb-2015 Formulations

Mean Concentration (mg/mL)

50.5

196

RSD (%)

0.61

0.57

Mean % of Target

101

98.2

11-Day Refrigerated Storage Resuspension Homogeneity Assessment of the 12-Feb-2015 Formulations

Mean Concentration (mg/mL)

51.3

201

RSD (%)

1.1

1.0

Mean % of Target

103

100

11-Day Room Temperature Storage Resuspension Homogeneity Assessment of the 12-Feb-2015 Formulations

Mean Concentration (mg/mL)

51.0

199

RSD (%)

0.37

0.56

Mean % of Target

102

99.7

 

Table 7.8.2/2: Results of Concentration Analyses

Date of Preparation

Mean Concentration, mg/mL (% of Target)

Group 2

(50 mg/mL)

Group 3

(100 mg/mL)

Group 4

(200 mg/mL)

12-Feb-2015

50.6 (101)

99.9 (99.9)

197 (98.6)

27-Feb-2015

51.3 (103)

104 (104)

197 (98.6)

Conclusions:
Under the test conditions, the NOAEL for maternal toxicity and embryo/fetal development is 1000 mg/kg bw/day in rats for the source and the target substances.
Executive summary:

In a prenatal developmental toxicity study performed according to OECD Guideline 414 and in compliance with GLP, the source substance was administered by oral (gavage) to groups of mated female Crl:CD(SD) rats (25/dose) at the dose levels of 0 (corn oil),250, 500, and 1000mg/kg bw/day from gestation days 2 through 19. The females were approximately 13 weeks of age at the initiation of dose administration. All animals were observed twice daily for mortality and moribundity. Clinical observations, body weights, and food consumption were recorded at appropriate intervals. On gestation day 20, a laparohysterectomy was performed on each female. The uteri, placentae, and ovaries were examined, and the numbers of fetuses, early and late resorptions, total implantations, and corpora lutea were recorded. Gravid uterine weights were recorded, and net body weights and net body weight changes were calculated. Selected organs were weighed. The fetuses were weighed, sexed, and examined for external, visceral, and skeletal malformations and developmental variations.

 

All females in the control, 250, 500, and 1000 mg/kg bw/day groups survived to the scheduled necropsy on gestation day 20. Test item-related clinical findings were limited to dose-dependent clear and red material around the mouth in all test item-treated groups approximately 1 hour following dose administration generally throughout the treatment period. These findings did not persist to the daily examinations (prior to dose administration) on the following day and were not considered to be adverse.

 

Test item-related slightly lower mean body weight gains were noted in the 250, 500, and 1000 mg/kg bw/day groups during gestation days 2-6. Mean body weight gains in the test item-treated groups were similar to the control group throughout the remainder of the treatment period (gestation days 6-19) and there were no corresponding effects on mean food consumption or mean absolute body weights. Therefore, the initial body weight decrements in the 250, 500, and 1000 mg/kg bw/day groups were not considered to be adverse. Mean maternal body weights, net body weights, net body weight gains, gravid uterine weights, and food consumption in the 250, 500, and 1000 mg/kg bw/day groups were unaffected by test item administration. No test item-related macroscopic findings or effects on brain, liver, thymus, and spleen weights were noted at any dosage level. Intrauterine growth and survival and fetal morphology in the 250, 500, and 1000 mg/kg bw/day groups were unaffected by maternal test item administration.

 

Based on the lack of adverse maternal or developmental toxicity observed at any dosage level, the NOAEL for maternal toxicity and embryo/fetal development is 1000 mg/kg bw/day in rats for the source and the target substances.

Effect on developmental toxicity: via oral route
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
1 000 mg/kg bw/day
Study duration:
subacute
Species:
rat
Quality of whole database:
The key study is GLP-compliant and of high quality (Klimisch score = 1 ).
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

There are no reproductive toxicity studies on the test material. A pre-natal / developmental toxicity study performed according to OECD 414 is available on a supporting substance, Cyclohexane, 1 -4 -dicarboxylic acid.

Study summary:

In a prenatal developmental toxicity study performed according to OECD Guideline 414 and in compliance with GLP, test substance was administered by oral (gavage) to groups of mated femaleCrl:CD(SD)rats (25/dose) at the dose levels of 0 (corn oil),250, 500, and 1000mg/kg bw/dayfrom gestation days 2 through 19.The females were approximately 13 weeks of age at the initiation of dose administration. All animals were observed twice daily for mortality and moribundity. Clinical observations, body weights, and food consumption were recorded at appropriate intervals. On gestation day 20, a laparohysterectomy was performed on each female. The uteri, placentae, and ovaries were examined, and the numbers of fetuses, early and late resorptions, total implantations, and corpora lutea were recorded. Gravid uterine weights were recorded, and net body weights and net body weight changes were calculated. Selected organs were weighed. The fetuses were weighed, sexed, and examined for external, visceral, and skeletal malformations and developmental variations.

 

All females in the control, 250, 500, and 1000 mg/kg bw/day groups survived to the scheduled necropsy on gestation day 20. Test item-related clinical findings were limited to dose-dependent clear and red material around the mouth in all test item-treated groups approximately 1 hour following dose administration generally throughout the treatment period. These findings did not persist to the daily examinations (prior to dose administration) on the following day and were not considered to be adverse.

 

Test item-related slightly lower mean body weight gains were noted in the 250, 500, and 1000 mg/kg bw/day groups during gestation days 2-6. Mean body weight gains in the test item-treated groups were similar to the control group throughout the remainder of the treatment period (gestation days 6-19) and there were no corresponding effects on mean food consumption or mean absolute body weights. Therefore, the initial body weight decrements in the 250, 500, and 1000 mg/kg bw/day groups were not considered to be adverse. Mean maternal body weights, net body weights, net body weight gains, gravid uterine weights, and food consumption in the 250, 500, and 1000 mg/kg bw/day groups were unaffected by test item administration. No test item-related macroscopic findings or effects on brain, liver, thymus, and spleen weights were noted at any dosage level. Intrauterine growth and survival and fetal morphology in the 250, 500, and 1000 mg/kg bw/day groups were unaffected by maternal test item administration.

 

Based on the lack of adverse maternal or developmental toxicity observed at any dosage level, the NOAEL for maternal toxicity and embryo/fetal development is 1000 mg/kg bw/day in rats for both the source and the target substances.

Justification for classification or non-classification

Harmonized classification:

The substance has no harmonized classification according to the regulation (EC) No. 1272/2008.

Self-classification:

The substance is not considered to have adverse effects on reproduction based on the results of an OECD 421 and an OECD 414 performed on two valid supporting substances. Classification of the substance for this endpoint is not justified in the absence of treatment-related effect according to the Regulation (EC) No. 1272/2008 (CLP).