4-(1,1-dimethylethyl)cyclohexyl acrylate

Toxicity to reproduction

Administrative data

Endpoint:

screening for reproductive / developmental toxicity

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2015

Report date:

2015

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

other: EPA, Health Effects Test Guidelines; OPPTS 870.3650: Combined Repeated Dose Toxicity Study With the Reproduction/Developmental Toxicity Screening Test (Jul 2000)

GLP compliance:

yes (incl. QA statement)

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

4-(1,1-dimethylethyl)cyclohexyl acrylate

EC Number:

282-104-8

EC Name:

4-(1,1-dimethylethyl)cyclohexyl acrylate

Cas Number:

84100-23-2

Molecular formula:

C13H22O2

IUPAC Name:

4-tert-butylcyclohexyl prop-2-enoate

Test material form:

other: liquit

Details on test material:

- Name of test material (as cited in study report): Laromer TBCH
- Physical state: liquid / colorless, clear
- Analytical purity: 92.7%
- Lot/batch No.: 140004P040

Test animals

Species:

rat

Strain:

Wistar

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories, Research Models and Services, Germany GmbH
- Age at study initiation: 10 - 11 weeks (males/females)
- Housing: individually (polycarbonate cages type III supplied by TECNIPLAST)
- Exceptions:
- During overnight matings, male and female mating partners were housed together in polycarbonate cages type III.
- Pregnant animals and their litters were housed together until PND 4 (end of lactation).
- Diet: ad libitum (ground Kliba maintenance diet mouse-rat “GLP”, meal, supplied by Provimi Kliba SA, Kaiseraugst, Switzerland)
- Water: ad libitum (dringing water)
- Acclimation period: at least 7 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-24°C
- Humidity (%): 30-70%
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

corn oil

Details on exposure:

PREPARATION OF DOSING SOLUTIONS:
The substance was applied as a solution. To prepare this solution, the appropriate amount of test substance was weighed out depending on the desired concentration. Then, corn oil was filled up to the desired volume, subsequently released with a magnetic stirrer. The test substance preparations were produced weekly and stored at room temperature.

VEHICLE
- Justification for use and choice of vehicle: In the context of toxicological studies the stability of the test substance in the vehicle corn oil has to be verified.
- Concentration in vehicle: 1.25 g/100 mL (50 mg/kg bw/day); 3.75 g/100 mL (150 mg/kg bw/day); 12.50 g/100 mL (500 mg/kg bw/day)
- Amount of vehicle: 4 mL/kg bw

Details on mating procedure:

In general, each of the male and female animals was mated overnight in a 1:1 ratio for a maximum of 2 weeks. Throughout the mating period, each female animal was paired with a predetermined male animal from the same test group.
The animals were paired by placing the female in the cage of the male mating partner from about 16.00 h until 07.00 - 09.00 h of the following morning. Deviations from the specified times were possible on weekends and public holidays and were reported in the raw data. A vaginal smear was prepared after each mating and examined for the presence of sperm. If sperm was detected, pairing of the animals was discontinued. The day on which sperm was detected was denoted gestation day (GD) 0 and the following day "GD 1".

Analytical verification of doses or concentrations:

no

Details on analytical verification of doses or concentrations:

In the context of toxicological studies the stability of the test substance in the vehicle corn oil has to be verified.
Concentration control analyses of the test substance preparations were performed in samples of all concentrations at the start of the administration period. Given that the test substance is completely miscible with corn oil, solutions were considered to be homogenous and no homogeneity analyses were carried out.

Duration of treatment / exposure:

males: 28 days
female: 57 days

Frequency of treatment:

daily

Details on study schedule:

- Dose selection rationale: Dose levels were selected by request of the sponsor.
- Route of administration: The oral route was selected since administration by gavage has been proven to be appropriate for the detection of a toxicological hazard.

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

0 mg/kg bw/day (actual dose received)

Doses / concentrations 2

Dose / conc.:

50 mg/kg bw/day (actual dose received)

Doses / concentrations 3

Dose / conc.:

150 mg/kg bw/day (actual dose received)

Doses / concentrations 4

Dose / conc.:

500 mg/kg bw/day (actual dose received)

No. of animals per sex per dose:

10

Control animals:

yes, concurrent vehicle

Examinations

Parental animals: Observations and examinations:

MORTALITY / CAGE SIDE OBSERVATIONS: Yes
- Time schedule: A cageside examination was conducted at least once daily for any signs of morbidity, pertinent behavioral changes and signs of overt toxicity.Abnormalities and changes were documented daily for each affected animal. The littering and lactation behavior of the dams was generally evaluated in the mornings in combination with the daily clinical inspection of the dams. Only particular findings (e.g. inability to deliver) were documented on an individual dam basis. On weekdays (except public holidays) the parturition behavior of the dams was inspected in the afternoons in addition to the evaluations in the mornings. The day of littering was considered the 24-hour period from about 15.00 h of one day until about 15.00 h of the following day.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Detailed clinical observations (DCO) were performed in all animals prior to the administration period and thereafter at weekly intervals. The findings were ranked according to the degree of severity, if applicable. The animals were transferred to a standard arena (50 × 37.5 cm with sides of 25 cm high). The following parameters were examined: abnormal behavior in handling, fur, skin, posture, salivation, respiration, activity/arousal level , tremors, convulsions, abnormal movements, gait abnormalities, lacrimation, palpebral closure, exophthalmos, assessment of the feces discharged during the examination (appearance/consistency), assessment of the urine discharged during the examination, pupil size

BODY WEIGHT: Yes
Body weight was determined before the start of the administration period in order to randomize the animals. During the administration period body weight was determined on study day 0 (start of the administration period) and thereafter once a week at the same time of the day (in the morning).
The body weight change of the animals was calculated from these results. The following exceptions are notable for the female animals:
- During the mating period the parental females were weighed on the day of positive evidence of sperm (GD 0) and on GD 7, 14 and 20.
- Females with litter were weighed on the day of parturition (PND 0) and on PND 4.
- Females without a litter and without positive evidence of sperm in the vaginal smear or waiting for necropsy were weighed weekly. These body weight data were solely used for the calculations of the dose volume.

FOOD CONSUMPTION:
Generally, food consumption was determined once a week for male and female parental animals, with the following exceptions:
- Food consumption was not determined during the mating period (male and female F0 animals).
- Food consumption of the F0 females with evidence of sperm was determined on GD 0-7, 7-14 and 14-20.
- Food consumption of F0 females, which gave birth to a litter was determined for PND 1-4.
Food consumption was not determined in females without positive evidence of sperm (during the mating period of dams used in parallel) and females without litter (during the lactation period of dams used in parallel) and in males after the premating period.

WATER CONSUMPTION:
Drinking water consumption was monitored by daily visual inspection of the water bottles for any changes in volume.

HAEMATOLOGY: Yes
- Time schedule for collection of blood: In the morning towards the end of tha administrative period.
- Anaesthetic used for blood collection: Yes (Isoflurane)
- Animals fasted: Yes
- How many animals: first 5 surviving parental males and the first 5 surviving females with litter (in order of delivery) per group

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: In the morning towards the end of tha administrative period.
- Animals fasted: Yes
- How many animals: first 5 surviving parental males and the first 5 surviving females with litter (in order of delivery) per group

URINALYSIS: Yes
- Time schedule for collection of urine: overnight towards the end of the administrative period.
- Metabolism cages used for collection of urine: Yes
- Animals fasted: Yes

NEUROBEHAVIOURAL EXAMINATION: Yes
A functional observational battery (FOB) was performed in the first five surviving male animals per test group and the first five surviving females with litter (in order of delivery) of all test groups at the end of the administration period
- Dose groups that were examined: all amimals
- Battery of functions tested:
Home cage observations:
The animals were observed in their closed home cages; during this period any disturbing activities (touching the cage or rack, noise) were avoided during these examinations in order not to influence the behavior of the animals.
Attention was paid to: Posture, tremors, convulsions, abnormal movements, impairment of gait
Open field observations:
The animals were transferred to a standard arena (50 × 50 cm with sides of 25 cm height) and observed for at least 2 minutes. The following parameters were examined: Behavior on removal from the cage, fur, skin, salivation, nose discharge, lacrimation, eyes/pupil size, posture, palpebral closure, respiration, tremors,
convulsions, abnormal movements/stereotypes, impairment of gait, activity/arousal level, feces excreted within 2 minutes (appearance/consistency), urine excreted within 2 minutes (amount/color, rearing within 2 minutes.
Sensory motor tests/ reflexes:
The animals were then removed from the open field and subjected to following sensory motor or reflex tests: 1. Reaction to an object being moved towards the face (approach response), touch sensitivity (touch response), vision (visual placing response), pupillary reflex, pinna reflex, audition (auditory startle response), coordination of movements (righting response), behavior during handling, vocalization, pain perception (tail pinch), grip strength of forelimbs and hindlimbs, landing foot-splay test.

Motor activity (MA) was also measured on the same day as the FOB was performed in the first five parental males and the first five surviving females with litter (in order of delivery) per group.

Sperm parameters (parental animals):

stages of spermatogenesis

Litter observations:

PARAMETERS EXAMINED
The following parameters were examined in F1 offspring:
- number and sex of pups
- stillbirths
- live births
- postnatal mortality
- presence of gross anomalies
- weight gain, physical or behavioural abnormalities

BODY WEIGHT
The pups were weighed on the day after birth (PND 1) and on PND 4.

CLINICAL OBSERVATIONS
The live pups were examined daily for clinical symptoms (including grossmorphological findings) during the clinical inspection of the dams.

STANDARDISATION OF LITTERS
Pups were scheduled sacrifice on PND 4.

Postmortem examinations (parental animals):

GROSS PATHOLOGY: Yes
All parental animals were sacrificed by decapitation under isoflurane anesthesia. The exsanguinated animals were necropsied and assessed by gross pathology, special attention being given to the reproductive organs.
Organ weights
- The following weights were determined in all animals sacrificed on schedule: Anesthetized animals, epididymides, tstes.
- The following weights were determined in 5 animals/sex and test group (females with litters, same animals as used for clinical pathology examinations): Adrenal glands, brain, heart, kidneys, liver,spleen, thymus.

The following organs or tissues of all parental animals were fixed in in 4% neutralbuffered formaldehyde or in modified Davidson’s solution: All gross lesions, adrenal glands, aorta, bone marrow (femur), brain, cecum, cervix, coagulating glands, colon, duodenum, eyes with optic nerve, esophagus, extraorbital lacrimal glands, epididymides (modified Davidson’s solution), femur with knee joint, heart, ileum, jejunum (with Peyer’s patches), kidneys, larynx, liver, lungs, lymph nodes (axillary and mesenteric), mammary gland (male and female), nose (nasal cavity), ovaries (modified Davidson’s solution), oviducts, pancreas, parathyroid glands, pharynx, pituitary gland, prostate gland, rectum, salivary glands (mandibular and sublingual), sciatic nerve, seminal vesicles, skeletal muscle, spinal cord (cervical, thoracic and lumbar cord), spleen, sternum with marrow, stomach (forestomach and glandular stomach), testes (modified Davidson’s solution), thymus, thyroid glands, trachea, urinary bladder, uterus (uteri of all cohabited female F0 parental animals were stained according to Salewski’s method), vagina.

Postmortem examinations (offspring):

All pups with scheduled sacrifice on PND 4 were sacrificed under isoflurane anesthesia with CO2. All pups were examined externally and eviscerated; their organs were assessed macroscopically.
All stillborn pups and all pups that died before PND 4 were examined externally, eviscerated and their organs were assessed macroscopically.
All pups without notable findings or abnormalities were discarded after their macroscopic evaluation. Animals with notable findings or abnormalities were evaluated on a case-by-case basis, depending on the type of finding noted.

Statistics:

Statistics of clinical pathology
Means, medians and standard deviations of each test group were calculated for several parameters.
Blood parameters: For parameters with bidirectional changes: Non-parametric one-way analysis using KRUSKAL-WALLIS test. If the resulting p-value was equal or less than 0.05, a pairwise comparison of each dose group with the control group was performed using WILCOXON-test (twosided) for the hypothesis of equal medians For parameters with unidirectional changes: Pairwise comparison of each dose group with the control group using the WILCOXON-test (one-sided) with Bonferroni-Holm adjustment for the hypothesis of equal medians
Urinalysis parameters (apart from pH, urine volume, specific gravity, color and turbidity): Pairwise comparison of each dose group with the control group using the WILCOXON-test (one-sided) for the hypothesis of equal medians.
Urine pH, volume, specific gravity, color and turbidity: Non-parametric one-way analysis using KRUSKAL-WALLIS test. If the resulting p-value was equal or less than 0.05, a pairwise comparison of each dose group with the control group was performed using WILCOXON-test (twosided) for the hypothesis of equal medians. Urine color and turbidity are not evaluated statistically.

Statistics of pathology
Weight parameters: Non-parametric one-way analysis using KRUSKAL-WALLIS test (two-sided). If the resulting pvalue was equal or less than 0.05, a pairwise comparison of each dose group with the control group was performed using WILCOXON-test (two-sided) for the equal medians

Reproductive indices:

Male reproduction data
The pairing partners, the number of mating days until vaginal sperm was detected in the female animals, and the gestational status of the females were recorded for F0 breeding pairs. For the males, mating and fertility indices were calculated for F1 litters.
Male mating index (%) = (number of males with confirmed mating* / number of males placed with females) x 100
*defined by a female with vaginal sperm or with implants in utero
number of males proving their fertility*
Male fertility index (%) = (number of males proving their fertility* / number of males placed with females) x 100
* defined by a female with implants in utero

Female reproduction and delivery data
The pairing partners, the number of mating days until vaginal sperm were detected and gestational status were recorded for F0 females. For the females, mating, fertility and gestation indices were calculated for F1 litters.
number of females mated*
Female mating index (%) = (number of females mated* / number of females placed with males) x 100
* defined as the number of females with vaginal sperm or with implants in utero
number of females pregnant*
Female fertility index (%) = (number of females pregnant* / number of females mated**) x 100
* defined as the number of females with implants in utero
** defined as the number of females with vaginal sperm or with implants in utero
Gestation index (%) = (number of females with live pups on the day of birth / number of females pregnant*) x 100
* defined as the number of females with implants in utero
number of liveborn pups at birth
Live birth index (%) = (number of liveborn pups at birth / total number of pups born) x 100
total number of pups born
The implantations were counted and the postimplantation loss (in %) was calculated.
Post implantation loss (%) = ((number of implantations - number of pups delivered) / number of implantations) x 100

Offspring viability indices:

Pup number and status at delivery
All pups delivered from the F0 parents (F1 litter) were examined as soon as possible on the day of birth to determine the total number of pups, the sex and the number of liveborn and stillborn pups in each litter. At the same time, the pups were also being examined for macroscopically evident changes. Pups, which died before this initial examination, were defined as stillborn pups.

Pup viability/mortality
The number and percentage of dead pups on the day of birth (PND 0) and of pups dying between PND 1-4 (lactation period) were determined. Pups which died accidentally or were sacrificed due to maternal death were not included in these calculations. The number of live pups/litter was calculated on the day after birth, and on lactation day 4.
Viability index (%) = (number of live pups on day 4 after birth / number of live pups on the day of birth) x 100

Sex ratio:
On the day of birth (PND 0) the sex of the pups was determined by observing the distance between the anus and the base of the genital tubercle; normally, the anogenital distance is considerably greater in male than in female pups. The sex of the pups was finally confirmed at necropsy. The sex ratio was calculated at day 0 and day 4 after birth.
Sex ratio = (number of live male or female pups on day 0/4 / number of live male and female pups on day 0/4) x 100

Results and discussion

Results: P0 (first parental generation)

General toxicity (P0)

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

Tonic-clonic convulsions were observed in 5 high dose female animals on different days during lactation period. Twitching (mostly slight, sometimes moderate) was observed during the entire administration period in 9 male and all female animals of test group 3 (500 mg/kg bw/d) from premating day 8 onwards on different study days (mostly 0-2 hours, partly 2-5 hours after treatment). The findings were assessed as treatement related and adverse.
Piloerection was observed in 2 male animals of test group 3 (500 mg/kg bw/d) from mating day 2 (study day 15) onwards on different study days. The finding was assessed as being related to treatment and adverse. A single male animal of test group 2 (150 mg/kg bw/d) showed piloerection just on mating day 2. In this case, a relation to treatment was not assumed because of the transient and short occurrence of the finding. Plough nose-first into bedding was observed shortly after treatment during the entire administration period (except gestation and lactation) on different study days in all males and 4 females of test group 3 (500 mg/kg bw/d) as well as in 6 males and 1 female of test group 2 (150 mg/kg bw/d). Salivation shortly after treatment, mostly slight, sometimes moderate, was observed in all phases of the study for all male and female animals of test group 3 (500 mg/kg bw/d) as well as for 7 male and 2 female animals of test group 2 (150 mg/kg bw/d). From the temporary, short appearance immediately after dosing it was concluded that plough nose-first into bedding and salivation were induced by a bad taste of the test substance or local affection of the upper digestive tract.

Mortality:

no mortality observed

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

Mean body weight of male animals of test group 3 (500 mg/kg bw/d) was significantly reduced on mating day 8 (-5%). Mean body weight change values in male animals of test group 3 (500 mg/kg bw/d) were significantly lower between premating days 0-7 (- 53%) and 0-13 (-41%). These changes were assessed to be related to treatment and adverse.
No significant changes were observed in female animals of test group 3 (500 mg/kg bw/d) as well as in male and female animals of test groups 1 and 2 (50 and 150 mg/kg bw/d).

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Description (incidence and severity):

During the premating period food consumption was significantly decreased in males of test group 3 (500 mg/kg bw/d) between study days 0-7 and 0-13 as well as in females of the same test group between study days 0-7, 7-13 and 0-13. The changes were assessed to be related to treatment. No comparable observation were made in any other test group of the study. No impairment of food consumption was observed for female animals of any test group during the gestation and lactation periods.

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

no effects observed

Haematological findings:

effects observed, non-treatment-related

Description (incidence and severity):

No treatment-related, adverse changes among hematological parameters were observed. In male rats of test group 3 (500 mg/kg bw/d) absolute neutrophil cell counts and in females of the same test group absolute monocyte cell counts were higher compared to controls. In males the absolute neutrophil counts were within the historical control range and in females the absolute monocyte counts were marginally above the range (males absolute neutrophils 0.70-1.35 Giga/L; females absolute monocytes 0.05-0.09 Giga/L; PART III, Supplement). No other changes in the differential blood cell counts occurred. Therefore, the change in males was regarded as incidental and not treatment-related, and the marginally higher monocyte counts in females were regarded as maybe treatment-related, but not adverse (ECETOC Technical Report No. 85, 2002).
In males of test group 2 (150 mg/kg bw/d) relative neutrophil cell counts were decreased and relative lymphocyte cell counts were increased. Both parameters were not dose-dependently changed and therefore this alteration was regarded as incidental and not treatment-related.

Clinical biochemistry findings:

effects observed, treatment-related

Description (incidence and severity):

The only treatment related effect were increased urea levels and decreased glucose levels in high dose males. These changes might be related to the alpha 2µ nephropathy observed in all dose groups, which does not represent a human relevant hazard.

In females of test groups 2 and 3 (150 and 500 mg/kg bw/day) inorganic phosphate and cholesterol levels were increased. However, cholesterol levels in both mentioned test groups and inorganic phosphate values at least in test group 2 were within the historical control ranges (cholesterol 1.13-1.77 mmol/L; inorganic phosphate 1.14-1.55 mmol/L; PART III, Supplement). Therefore, the changes in the mentioned test groups were regarded as incidental and not treatment-related. Higher inorganic phosphate levels in females of test group 3 (500 mg/kg bw/dây) were the only altered parameter in these individuals and therefore this increase was regarded as maybe treatment-related but not adverse (ECETOC Technical Report No. 85, 2002). In males of test groups 1 and 2 (50 and 150 mg/kg bw/day) total bile acid levels were lower compared to controls, but this change was not dose-dependent. Therefore, it was regarded as incidental and not treatment-related.

Urinalysis findings:

effects observed, treatment-related

Description (incidence and severity):

No treatment-related, adverse changes among urinalysis parameters were observed.

In males of test group 3 (500 mg/kg bw/day) urine pH value was lower and in females of the same test group urine volume was higher compared to controls. Both parameter changes without any other changes in the kidneys were regarded as treatment-related but not adverse.
In males of test groups 1, 2 and 3 (50, 150 and 500 mg/kg bw/day) higher incidences of transitional epithelial cells and granulated and epithelial casts were found in the urine sediment. This observation correlates well with the histopathological finding of alpha 2µ-Globulinurie, which was regarded as a rat specific effect with no relevance to humans (Hard et al., 1993).

Behaviour (functional findings):

effects observed, treatment-related

Description (incidence and severity):

Home cage observations: Twitching was observed in 2 females (Nos. 137 and 138) of test group 3 (500 mg/kg bw/day).
Open field observations: Twitching was observed in 2 females (Nos. 137 and 138) of test group 3 (500 mg/kg bw/day).
Sensorimotor tests/reflexes: No test substance-related effects were observed.
Quantitative Parameters: No test substance-related effects were observed.
Motor activity measurement: A significant deviation concerning the overall motor activity (summation of all intervals) was observed in female animals of test group 1 (50 mg/kg bw/day) in comparison to the concurrent control group. Regarding single intervals in male animals of test group 1 (50 mg/kg bw/day) a significantly reduced activity was observed in interval 6 as well as in female animals of test group 1 (50 mg/kg bw/day) in interval 11. As no other single intervals as well as the overall motor activity showed significant deviations to the control values and a dose-response relationship was not observed, the finding was assessed to be incidental and not related to treatment.

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

Treatment-related findings were observed in the kidneys of male animals. The number and pattern of eosinophilic droplets in the proximal convoluted tubules was highly discerned in the H&E stained tissues. Thus, a dose-dependent increase of eosinophilic droplets was noted. This finding was characterized not only by an increase in the number of droplets in the tubular epithelium cells but also by an increase in the number of affected tubules. With increasing dose, the form of the eosinophilic droplets changed to more angular and crystal-shaped eosinophilic aggregates. Concomitantly, tubular degeneration and regeneration in the cortex and outer stripe of the outer medulla (OSOM) and granular casts at the junction of the OSOM and the inner stripe of the outer medulla (ISOM) accompanied this process. The immunostaining with an antibody to α2µ-globulin correlated partially with the distribution pattern of eosinophilic droplets. All other findings occurred either individually or were biologically equally distributed over control and treatment groups. They were considered to be incidental or spontaneous in origin and without any relation to treatment.

Histopathological findings: neoplastic:

not examined

Reproductive function / performance (P0)

Reproductive performance:

no effects observed

Description (incidence and severity):

The male mating index was 90% in control group (0 mg/kg bw/day) and 100% in test groups 1-3 (50, 150 and 500 mg/kg bw/day). The male fertility index was 80% in control and high dose rats, 90% in the low dose and 70% in the mid dose. These values reflected the normal range of biological variation inherent in the strain of rats used for this study. The non-pregnant females as well as their male mating partners did not show alterations in the histopathologic examination that could have impaired their fertility.

The female mating index calculated after the mating period for F1 litter was 90% in control group (0 mg/kg bw/day) and 100% in test groups 1-3 (50, 150 and 500 mg/kg bw/day). The mean duration until sperm was detected varied between 1.6 and 3.1 days without relation to dose and within the normal range for this strain of rats. Gestation length was similar for all dose groups.

Effect levels (P0)

1

Key result

Dose descriptor:

NOAEL

Remarks:

(parental systemic toxicity)

Effect level:

150 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

clinical signs

body weight and weight gain

Remarks on result:

other: tubular damage in the kidneys of male animals of all test groups as a consequence of an α2 µ nephropathy does not represent a risk for humans

2

Key result

Dose descriptor:

NOAEL

Remarks:

(parental repoductive toxicity- fertility)

Effect level:

500 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: no effects observed

Target system / organ toxicity (P0)

Key result

Critical effects observed:

no

Results: F1 generation

General toxicity (F1)

Clinical signs:

no effects observed

Mortality / viability:

mortality observed, non-treatment-related

Description (incidence and severity):

The viability index indicating pup mortality during lactation (PND 0-4) was 98.1% in control animals, 100% in the low and mid dose and 96.7% in the high dose. These findings reflected the normal range of biological variation inherent in the strain of rats used for this study as all respective values were within the range of the historical control data. Two pups in the control group and 4 pups in the high dose were cannibalized. One pup in the high dose group was stillborn. A relation to treatment was excluded.

Body weight and weight changes:

no effects observed

Description (incidence and severity):

Mean pup body weights/pup body weight changes of all pups in all test groups were comparable to the control group. One male runt each was found in the control and the low dose. Two female runts occurred in the mid dose. One male and one female runt were found in the high dose.

Gross pathological findings:

no effects observed

Other effects:

no effects observed

Description (incidence and severity):

The sex distribution and sex ratios of live F1 pups on the day of birth and PND 4 did not show substantial differences between the control and the test substance-treated groups.

Effect levels (F1)

Key result

Dose descriptor:

NOAEL

Remarks:

(developmental toxicity)

Generation:

F1

Effect level:

500 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: no effects observed

Target system / organ toxicity (F1)

Key result

Critical effects observed:

no

Overall reproductive toxicity

Key result

Reproductive effects observed:

no