Registration Dossier

Toxicological information

Toxicity to reproduction

Currently viewing:

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:
2018
Report Date:
2018

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)
GLP compliance:
yes (incl. certificate)

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
liquid

Test animals

Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Chales River Laboratories, Research Models and Services, Germany GmbH/ Charles River Laboratories, France
- Age at study initiation: 11 weeks (male animals); 10 weeks (female animlas)
- Housing: During the pretreatment period of the study, the rats were housed together (up to 5 animals per sex and cage) in Polysulfonate cages Typ 2000P (H-Temp) supplied by TECHNIPLAST, Ho
henpeißenberg, Germany. During the study period, the rats were housed individually in Polycarbonate cages type III supplied by TECHNIPLAST, Hohenpeißenberg, Germany and Becker & Co., Castrop-Rauxel, Germany, with the following 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 13 in Polycarbonate cages type III.
- Diet (e.g. ad libitum): ad libitum (ground Kliba maintenance diet mouse/rat “GLP” meal, supplied by Provimi Kliba SA, Kaiseraugst, Switzerland)
- Water (e.g. ad libitum): ad libitum

ENVIRONMENTAL CONDITIONS
- Temperature (°C):20-24
- Humidity (%): 30-70
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
water
Details on exposure:
The test substance solutions in drinking water were prepared in intervals, which took into account the analytical results of the stability verification. For the preparation of the administration solutions the test substance was weighed in a graduated flask depending on the dose group, topped up with drinking water and intensely mixed with a magnetic stirrer.
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 dose group. If sperm was detected, pairing of the animals was discontinued. The day on which sperm were detected was denoted " gestation day (GD) 0" and the following day "gestation day (GD) 1"
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
The stability of the test substance in drinking water for a period of 7 days at room temperature were carried out.
Duration of treatment / exposure:
After the acclimatization period, the test substance was administered to the parental animals orally by gavage.
Frequency of treatment:
once daily at approximately the same time in the morning
Doses / concentrationsopen allclose all
Dose / conc.:
0 mg/kg bw/day (nominal)
Dose / conc.:
15 mg/kg bw/day (nominal)
Dose / conc.:
50 mg/kg bw/day (nominal)
Dose / conc.:
150 mg/kg bw/day (nominal)
No. of animals per sex per dose:
10
Control animals:
yes, concurrent vehicle

Examinations

Parental animals: Observations and examinations:
Mortality
A check for moribund and dead animals was made twice daily on working days and once daily on Saturdays, Sundays and public holidays. If animals were in a moribund state, they were sacrificed and necropsied.

Clinical observations
A cageside examination was conducted at least once daily for any signs of morbidity,
pertinent behavioral changes and/or signs of overt toxicity. All animals were checked daily for any abnormal clinical signs before the administration as well as within 2 hours and within 5 hours after the administration. Abnormalities and changes were documented for each animal.

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 after the 2nd premating week (male parental animals) and during the mating period (male and female parental animals). Food consumption of the females with evidence of sperm was determined for GD 0-7, 7-14 and 14-20. Food consumption of the females which gave birth to a litter was determined for PNDs
1-4, 4-7, 7-10 and 10-13. Food consumption was not determined in females without positive evidence of sperm during the mating and the gestation period and in females without litter during the lactation period.

Body weight data
In general, the body weight of the male and female parental animals was determined once a week at the same time of the day (in the morning) until sacrifice.
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, 7, 10 and 13.
Females without positive evidence of sperm, without litter or waiting for necropsy, were weighed weekly. These body weight data were solely used for the calculations of the dose volume;

Detailed clinical observations
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.

Functional observational battery
A functional observational battery (FOB) was performed in the first five male and the first 5 female animals with litter per group (in order of delivery) at the end of the administration period starting at about 10.00 h. The FOB started in a randomized sequence with passive observations without disturbing the animals, followed by removal from the home cage, open field observations in a standard arena and sensorimotor tests as well as reflex tests. The findings were ranked according to the degree of severity, if applicable. The observations were performed at random.

Home cage observations:
The animals were observed in their closed home cages (for a short period: about 10-30 seconds); 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, gait, other findings

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, nasal discharge, lacrimation, eyes/pupil size, posture, palpebral closure, respiration, tremors, convulsions, abnormal movements/stereotypes, gait, activity/arousal level, feces excreted within 2 minutes (appearance/ consistency), urine excreted within 2 minutes (amount/color), rearing within 2 minutes, other findings

Sensory motor tests/ reflexes:
The animals were then removed from the open field and subjected to following sensory motor or reflex tests: 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), other findings, grip strength of forelimbs, grip strength of hindlimbs, landing foot-splay test

Motor activity assessment
The Measurement of motor activity (MA) was measured at the end of the administration period in the first 5 surviving parental males and the first 5 surviving females with litter (in order of delivery) per group. Motor activity (MA) was measured from 14:00 h onwards on the same day as the FOB was performed. The examinations were performed using the TSE Labmaster System supplied by TSE Systems GmbH, Bad Homburg, Germany. For the duration of the measurement the animals were placed in new clean polycarbonate cages with a small amount of bedding. Eighteen beams were allocated per cage. The number of beam interrupts were counted over 12 intervals of 5 minutes per interval. The sequence in which the animals were placed into the cages was selected at random. On account of the time needed to place the animals in the cages, the starting time was "staggered" for each animal. The measurement period began when the 1st beam was interrupted and was finished exactly 1 hour later. No food or water was offered to the animals during these measurements and the measurement room was darkened after the transfer of the last animal.

Clinical Pathology
In the morning blood was taken from the retro-bulbar venous plexus from fasted animals. The animals were anaesthetized using isoflurane. The blood sampling procedure and subsequent analysis of blood and serum samples were carried out in a randomized sequence. The assays of blood and serum parameters were performed under internal laboratory quality control conditions with reference controls to assure reliable test results. The results of clinical pathology examinations were expressed in International System (SI) units. The parameters listed below were examined in the first 5 surviving parental males per group at termination and in the first 5 females with litters (in order of delivery) per group at PND 14.

Hematology
The following parameters were determined in blood with EDTA-K3 as anticoagulant using a particle counter (Advia 120 model; Bayer, Fernwald, Germany): Leukocyte count, Erythrocyte count, Hemoglobin, Hematocrit, mean corpuscular volume, mean corpuscular Hemoglobin, mean corpuscular Hemoglobin concentration, Platelet count, Differential blood, Reticulocytes, Prothrombin time.

Clinical chemistry
Parameters: Alanine aminotransferase (ALT) (L-alanine: 2-oxoglutarate aminotransferase), Aspartate aminotransferase (AST) (L-aspartate: 2-oxoglutarate aminotransferase), Alkaline phosphatase (ALP) (orthophosphoric acid monoester phosphohydrolase), γ-Glutamyltransferase (GGT) (γ -glutamyl) peptide: aminoacid-γ- glutamyl-transferase, sodium, potassium, chloride, inorganic phosphate, calcium, urea, creatinine, glucose, total bilirubin, total protein, albumin, globulins, triglycerides, choloesterol, bile acids.

Thyroid Hormones
Blood samples were taken from all surplus pups or 2 preferably female pups per litter at PND 4 as well as one male and one female pup per litter at PND 13 by decapitation under isoflurane anesthesia. Additionally, blood samples from all dams at PND 14 and all males at termination were taken by puncturing the retrobulbar venous plexus under isoflurane anesthesia. The adults were fastened before the blood sampling.
All generated serum samples were frozen at -80°C until measurement.
Blood samples from the adult males and the PND 13 pups were assessed for serum levels for thyroid hormones (T4).
T4 ELISA was measured with a Sunrise MTP-reader, Tecan AG, Maennedorf, Switzerland, and evaluated with the Magellan-Software of the instrument producer.
Oestrous cyclicity (parental animals):
For all females in a pool of 50 animals, estrous cycle normality was evaluated before randomization (the estrous cycle data of these individuals are not reported and can be found in the raw data). For a minimum of 2 weeks prior to mating, estrous cycle length and normality was evaluated by daily analysis of vaginal smears for all F0 female parental rats. Determination was continued throughout the pairing period until the female exhibited evidence of copulation. At necropsy, one additional vaginal smear was examined to determine the stage of estrous cycle for each F0 female with scheduled sacrifice.
Litter observations:
Litter observation:
Litter data
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
In general, a check was made for any dead or moribund pups twice daily on workdays (once in the morning and once in the afternoon) or as a rule, only in the morning on Saturdays, Sundays or public holidays. Dead pups were evaluated by the methods, which are described in detail in “Pup Necropsy observations“. The number and percentage of dead pups on the day of birth (PND 0) and of pups dying
between PNDs 1-4, 5-7 and 8-13 were determined. Pups, which died accidentally or had to be sacrificed due to maternal death, were not included in these calculations. The number of live pups/litter was calculated on the day of birth (PND 0), and on lactation days 4, 7 and 13.

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. Later, during the course of lactation, this initial sex determination was followed up by surveying the external appearance of the anogenital region and the mammary line. The sex of the pups was finally confirmed at necropsy.

Pup clinical observations
The live pups were examined daily for clinical symptoms (including gross-morphological findings) during the clinical inspection of the dams.

Pup body weight data
The pups were weighed on the day after birth (PND 1) as well as on PND 4 (before
standardization), 7 and 13. Pups' body weight change was calculated from these results. The individual weights were always determined at about the same time of the day (in the morning). In the summary tables pup body weights and pup body weight change are listed for males, females and males + females. „Runts“ were defined on the basis of the body weights on PND 1. "Runts" are pups that weigh less than 75% of the mean weight of the respective control pups.
Postmortem examinations (parental animals):
Necropsy
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, Ovaries, Prostate, Seminal vesicles with coagulating glands, Testes, Thyroid glands (fixed) ,Uterus (with cervix).

The following weights were determined in 5 animals per sex/test group sacrificed on schedule (females with litters only, same animals as used for clinical pathological examinations): Adrenal glands, Brain, Heart, Kidneys, Liver, Spleen, Thymus.

Organ/tissue fixation
Parental animals
The following organs or tissues of all parental animals were fixed in in 4% neutral-buffered formaldehyde solution or in modified Davidson`s solution: All gross lesions, Adrenal glands, Aorta, Bone marrow (femur), Brain, Cecum, Cervix, Coagulating glands, Colon, Duodenum, Epididymides (modified Davidson`s solution), Esophagus, Extraorbital lacrimal glands, Eyes with optic nerve (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 apparently nonpregnant animals or empty uterus horns were stained according to Salewski E, 1964), Vagina.

Histopathology
Fixation was followed by histotechnical processing, examination by light microscopy and assessment of findings.
All gross lesion, Adrenal glands, Brain, Cecum, Cervix, Coagulating glands, Colon, Duodenum, Epididymides, Eyes with optic nerve, Heart, Ileum, Jejunum, Kidneys, Liver, Lungs, Lymph nodes (axillary and mesenteric), Ovaries, Oviducts, Prostate gland, Peyer`s patches, Rectum, Sciatic nerve, Seminal vesicles, Skeletal muscle, Spinal cord (cervical, thoracic, lumbar), Spleen, Sternum with marrow, Stomach (forestomach and glandular stomach), Testes, Thymus, Thyroid , glands, Trachea, Urinary bladder, Uterus, Vagina.
Postmortem examinations (offspring):
On PND 13, one selected male and one female pup per litter was sacrificed under isoflurane anesthesia by decapitation. Blood was sampled for determination of thyroid hormone concentrations. Thyroid glands/parathyroid glands were fixed in neutral buffered 4% formaldehyde solution and were transferred to the Pathology Laboratory for possible further processing.
Statistics:
Non-parametric one-way analysis using KRUSKAL-WALLIS test (two-sided). 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 (two-sided) for the equal medians.
Reproductive indices:
The following indeces were determined: mating and fertility indes for both males and females, gestation index, live birth index and postimplantation loss for female.
Offspring viability indices:
The following indeces were determined: viability index, survival index, sex ration, anogenital index.

Results and discussion

Results: P0 (first parental generation)

General toxicity (P0)

Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
Most animals of test group 3 as well as several animals of test group 2 (150 and 50 mg/kg bw/d) show ed salivation after treatment (grade: slight to severe) at some occasions during the study period. In female animals the incidence was generally higher during gestation and lactation periods, affecting most test group 3 and several test group 2 females. These observations were considered to be treatment-related.
No further clinical signs or changes of general behavior, which may be attributed to the test substance, were detected in any of the male and female F0 parental animals in any of the groups. Spontaneous findings were seen in one high-dose male (No. 33) which showed red discharge in the right eyeduring mating days 2 - 4, 8 - 11 and postmating days 0 – 1 as well as one sperm positive mid-dose female (No. 121) which did not deliver F1 pups.
Mortality:
no mortality observed
Body weight and weight changes:
effects observed, non-treatment-related
Description (incidence and severity):
Mean body weights and body weight change of all male and female F0 generation parental animals in all test substance-treated groups showed no significant difference to the concurrent control during the entire study.
The statistically significantly lower body weight change in the mid-dose females during PND 7 - 10 was considered as spontaneous in nature and not as treatment-related.
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
Food consumption of the F0 males in all dose groups (15, 50 and 150 mg/kg bw/d) as well as low and mid-dose females was not influenced by the treatment throughout the study.
Food consumption of the high-dose F0 females was statistically significantly below the concurrent control values during the entire premating period (up to 7%) while it remained unchanged during gestation and lactation periods.
Haematological findings:
effects observed, non-treatment-related
Description (incidence and severity):
In males of test group 3 (150 mg/kg bw/d) prothrombin time (Hepatoquick’s test, HQT) was significant ly reduced, but the mean was within the historical control range (males, HQT 37.4 - 40.9 sec). Therefore, this alteration was regarded as incidental and not treatment-related.
Clinical biochemistry findings:
effects observed, non-treatment-related
Description (incidence and severity):
In males of test groups 1 and 2 (15 and 50 mg/kg bw/d) cholesterol values were significantly changed (test group 1 lower, test group 2 higher) compared to controls, but the alteration was not dose-dependent. Therefore, this alteration was regarded as incidental and not treatmentrelated.
Behaviour (functional findings):
effects observed, non-treatment-related
Description (incidence and severity):
Functional observational battery (FOB)
Home cage observations:
No test substance-related or spontaneous findings were observed in male and female animals of all test groups during the home cage observation.
Open field observations: Two male animals of dose group 3 (Nos. 33 and 34 - 150 mg/kg bw/d) showed slight (area around the
mouth was moist) and severe (mouth very wet, wet paws) salivation, respectively. All other male and female animals of all dose groups (15, 50 and 150 mg/kg bw/d) did not show any abnormalities.
Sensorimotor tests/reflexes: There were no test substance-related findings in male and female animals of all test groups.
Quantitative Parameters: No test substance-related impaired parameters were observed in male and female animals of all test
groups. The statistically significantly higher value of the landing foot splay test in females of dose group 2 was considered as spontaneous in nature and not treatment related.
Motor activity measurement (MA)
No treatment-related changes of motor activity data was observed in all male and female animals of all dose groups (15, 50 and 150 mg/kg bw/d) in comparison to the concurrent control group. Overall activity levels and habituation to the test environment corresponded to the age of these animals, if usual biological variation inherent in the strain of rats used for this experiment was considered.
The isolated statistically significantly decreased numbers of beam interrupts in the males of dose groups 1 and 2 during interval 12 were not related to the dose and did not influence the overall activity levels and habituation. Thus, they were not considered to be related to the test substance.
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
Treatment-related findings were observed in forestomach and duodenum of male and female animals of test groups 2 and 3.
Duodenum: In the duodenum, a thickening of the mucosa characterized by an increase in villus height, was observed, which correlated to the macroscopic finding “dilation”. One female animal of test group 3 showed a focal hyperplasia of the duodenal mucosa.
Forestomach: Diffuse squamous hyperplasia (correlating with the macroscopic findings “margo plicatus, thickened”) and the presence of erosion/ulcer (correlating often with the macroscopic finding “focus”) were noted in the forestomach of male and female animals.
No treatment-related findings were noted in the glandular stomach and no correlate was found for the macroscopically observed discoloration in two test group 3 male animals. 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.The morphology of the testicular interstitium and the stages of spermatogenesis in the testes of males of the high dose (150 ppm) were comparable to those of the controls.

Reproductive function / performance (P0)

Reproductive function: oestrous cycle:
no effects observed
Description (incidence and severity):
Estrous cycle data, generated during the last 2 weeks prior to mating for the F1 litter, revealed regular cycles in the females of all test groups 0 - 3. The mean estrous cycle duration was similar: 4.0 days in test groups 0 - 3, respectively.
Reproductive performance:
no effects observed

Details on results (P0)

Thyroid hormones:
In the F0 parental males of test groups 2 and 3 (50 and 150 mg/kg bw/d) T4 values were significantly higher compared to controls. However, the means were within the historical control range (males T4 44.87-88.29 nmol/L). Additionally, neither a change of thyroid weight nor any histopathological findings in the thyroids were noted. Therefore, the higher T4 values in parental males were regarded as incidental and not treatment-related.
In male and female pups at PND13 (test groups 11, 12 and 13; 15, 50 and 150 mg/kg bw/d), no treatment-related alterations of T4 levels were observed.

Male reproduction data:
For all F0 parental males, which were placed with females to generate F1 pups, copulation was confirmed. Thus, the male mating index was 100% in test groups 0 - 3. Fertility was proven for most of the F0 parental males within the scheduled mating interval for
F1 litter.
One mid-dose male (50 mg/kg bw/d - No. 21) did not generate pregnancy.
Thus, the male fertility index ranged between 90% and 100% without showing any relation to dosing. This reflects the normal range of biological variation inherent in the strain of rats used for this study.

Female reproduction and delivery data:
The female mating index calculated after the mating period for F1 litter was 100% in all test groups. The mean duration until copulation was confirmed (GD 0) varied between 2.1 and 5.3 days without showing a statistically significant difference. The reason for the higher average in test group 3 were 3 mating pairs with copulation confirmed after 12 or 14 days of pairing, respectively. All 3 affected females had normal estrous cycles and normal pregnancies, and no findings were noted in the reproductive organs of the affected males and females. The very extensive period before copulation fits best to artificial pseudo-pregnancies provoked by the daily manipulations during vaginal lavage. None of this is considered to be treatment-related.
All female rats delivered pups or had implants in utero with the following exceptions: Mid-dose female No. 121 (mated with male No. 21) did not become pregnant.
The fertility index varied between 90% in test group 2 and 100% in the control and test groups 1 and 3. These values reflect the normal range of biological variation inherent in the strain of rats used for this study.
The non-pregnant female had no relevant gross lesions or microscopic findings.
The mean duration of gestation values varied between 21.9 (test group 1), 22.2 (control and test group 2) and 22.4 (test group 3), not indicating any influence by the test substance.
Implantation was not affected by the treatment since the mean number of implantation sites was comparable between all test substance-treated groups and the control, taking normal biological variation into account (12.8 / 13.0 / 13.2 and 14.3 implants/dam in test groups 0 - 3, respectively). Furthermore, there were no indications for test substance-induced intrauterine embryo-/fetolethality since the postimplantation loss did not show any significant differences between the groups, and the mean number of F1 pups delivered per dam remained unaffected (12.1 / 12.6 / 12.6 and 13.9 pups/dam in test groups 0 - 3, respectively).
The rate of liveborn pups was also not affected by the test substance, as indicated by live birth indices of 99.2% / 100% / 100% and 99.3% in test groups 0 - 3, respectively. Moreover, the number of stillborn pups was not significantly different between the test groups.
Thus, the test substance Hydroxypropylacrylate did not adversely affect reproduction and delivery of the F0 generation parental animals.

Effect levels (P0)

open allclose all
Dose descriptor:
NOAEL
Remarks:
fertility and reproductive performance
Effect level:
150 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: No adverse effects observed.
Dose descriptor:
NOAEL
Remarks:
developmental toxicity
Effect level:
150 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
female
Basis for effect level:
other: No adverse effects observed.
Dose descriptor:
NOAEL
Remarks:
general, systemic toxicity
Effect level:
150 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: No adverse effects observed.
Dose descriptor:
NOAEL
Remarks:
local effects
Effect level:
15 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
female
Basis for effect level:
other: irritation in forestomach and duodenum

Target system / organ toxicity (P0)

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):
PND 0: 1 male pup died in test group 0 and 3
PND 1-4: 1 male and 1 female pup died in test group 1; 2 male pups died in test group 2
Body weight and weight changes:
effects observed, non-treatment-related
Description (incidence and severity):
No test substance-related influence on body weights and body weight change values of F1 pups were noted in test groups 1 - 3 (15, 50 and 150 mg/kg bw/d).
One female runt was seen in the control, one male and two female runts were seen in test group 1, three male and two female runts were seen in test group 2 and one male and two female runts were seen in test group 3. These are spontaneous findings unrelated to the treatment.
Gross pathological findings:
effects observed, non-treatment-related
Description (incidence and severity):
A few pups showed spontaneous findings at gross necropsy, such as post mortem autolysis, discolored testis (left, red) and dilated renal pelvis.
These findings occurred without any relation to dosing and/or can be found in the historical control data at comparable or even higher incidences. Thus, all these findings were not considered to be associated to the test substance.

Details on results (F1)

Litter data
Pup number and status at delivery
The mean number of delivered F1 pups per dam and the rates of liveborn, stillborn, found dead and cannibalized F1 pups were evenly distributed about the test groups. The respective values reflect the normal range of biological variation inherent in the strain used in this study.
Pup viability
The viability index indicating pup mortality during early lactation (PND 0 - 4) varied between 100%/ 98.6% /97.9% and 100% in test groups 0 - 3, respectively. The survival index indicating pup mortality until mid-lactation (PND 4 - 13) was 100% in all test groups.
Sex ratio
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; slight differences were regarded to be spontaneous in nature.
Anogenital distance/anogenital index
No test substance-related effects were noted on anogenital distance or anogenital index in all treated F1 offspring (test groups 1 - 3 [15, 50 and 150 mg/kg bw/d]).
Nipple/ areola anlagen
The apparent number and percentage of male pups having areolae was not influenced by the test substance when examined on PND 13.

Effect levels (F1)

Dose descriptor:
NOAEL
Generation:
F1
Effect level:
150 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: No adverse effects observed.

Target system / organ toxicity (F1)

Critical effects observed:
no

Overall reproductive toxicity

Reproductive effects observed:
no

Applicant's summary and conclusion

Conclusions:
Under the conditions of the present OECD 422 combined repeated dose toxicity study with the reproductive/developmental screening test in Wistar rats, the NOAEL for fertility and reproductive performance was 150 mg/kg bw/d for the F0 parental rats. The NOAEL for developmental toxicity in the F1 progeny was 150 mg/kg bw/d.