Registration Dossier

Administrative data

Endpoint:
screening for reproductive / developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
15 June 2017 - 22 August 2017
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
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)
Limit test:
no
Justification for study design:
SPECIFICATION OF STUDY DESIGN FOR EXTENDED ONE-GENERATION REPRODUCTION TOXICITY STUDY WITH JUSTIFICATIONS [please address all points below]:

- Premating exposure duration for parental (P0) animals : 2 weeks
- Basis for dose level selection : preliminary tox studies
- Inclusion/exclusion of extension of Cohort 1B : not specified
- Termination time for F2 : not specified
- Inclusion/exclusion of developmental neurotoxicity Cohorts 2A and 2B : not examined
- Inclusion/exclusion of developmental immunotoxicity Cohort 3 : not examined
- Route of administration : oral gavage
- Other considerations, e.g. on choice of species, strain, vehicle and number of animals [if applicable]: A sufficient number of male and female Wistar Han™:RccHan™:WIST strain rats were obtained from Envigo RMS (UK) Limited, Blackthorn, Bicester, Oxon, UK. On receipt the animals were examined for signs of ill-health or injury. The animals were acclimatized for nineteen days during which time their health status was assessed. Following the day of arrival, vaginal smears were performed for all females throughout the acclimatization period. Any female considered not to be showing appropriate estrous cycling activity was excluded from treatment groups at least five days before the start of treatment. A total of ninety six animals (forty eight males and forty eight females) were accepted into the study. At the start of treatment the males weighed 280 to 355g and were approximately eleven weeks old. The females weighed 199 to 231g, and were approximately fourteen weeks old.

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
liquid: viscous
Specific details on test material used for the study:
Physical State/Appearance: Clear colorless viscous liquid
Purity: UVCB, 100%
Batch Number: WA 1508

Test animals

Species:
rat
Strain:
Wistar
Details on species / strain selection:
The rat was selected for this study as it is a readily available rodent species historically used in safety evaluation studies and is acceptable to appropriate regulatory authorities.
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Envigo RMS (UK) Limited, Blackthorn, Bicester, Oxon, UK
- Females (if applicable) nulliparous and non-pregnant: not specified
- Age at study initiation: At the start of treatment the males weighed 280 to 355g and were approximately eleven weeks old. The females weighed 199 to 231g, and were approximately fourteen weeks old.
- Weight at study initiation: At the start of treatment the males weighed 280 to 355g and were approximately eleven weeks old. The females weighed 199 to 231g, and were approximately fourteen weeks old.
- Fasting period before study: not specified
- Housing: Initially, all animals were housed in groups of three in solid floor polypropylene cages with stainless steel mesh lids and softwood flake bedding (Datesand Ltd., Cheshire, UK). During the pairing phase, animals were transferred to polypropylene grid floor cages suspended over trays lined with absorbent paper on a one male: one female basis within each dose group. Following evidence of successful mating, the males were returned to their original cages. Mated females were housed individually during gestation and lactation in solid floor polypropylene cages with stainless steel mesh lids and softwood flakes.
- Use of restrainers for preventing ingestion (if dermal): yes/no
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: 19 days
ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3 °C
- Humidity (%): 50 ± 20%
- Air changes (per hr): at least fifteen air changes per hour
- Photoperiod (hrs dark / hrs light): twelve hours continuous light and twelve hours darkness

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
polyethylene glycol
Details on exposure:
- PREPARATION OF DOSING SOLUTIONS:
For the purpose of this study the test item was prepared at the appropriate concentrations as a solution in Polyethylene glycol 400. The stability and homogeneity of the test item formulations were determined by Envigo Research Limited, Shardlow, UK, Analytical Services. Results showed the formulations to be stable for at least eleven days. Formulations were prepared daily for the first week of treatment (as stability had yet to be confirmed), formulations were then prepared weekly once eleven day stability data had been determined and stored at approximately 4 ºC in the dark.
Samples of test item formulation were taken on four occasions and analyzed for concentration of 1,3-Benzenedimethanamine, reaction products with glycidyl tolyl ether at Envigo Research Limited, Shardlow, UK, Analytical Services. The method used for analysis of formulations and the results obtained are given in Annex 2. The results indicate that the prepared formulations were within ± 10% of the nominal concentration, with the exception of Analysis 1 (daily preparation).

- DIET PREPARATION
The animals were allowed free access to food and water. A pelleted diet (Rodent 2018C Teklad Global Certified Diet, Envigo RMS (UK) Limited, Oxon, UK.) was used. Certificates of analysis of the batches of diet used are given in Annex 6. Mains drinking water was supplied from polycarbonate bottles attached to the cage. Environmental enrichment was provided in the form of wooden chew blocks and cardboard fun tunnels (Datesand Ltd., Cheshire, UK) except for paired animals and mated females during gestation and lactation. Mated females were also given softwood flakes, as bedding, throughout gestation and lactation. The diet, drinking water, bedding and environmental enrichment was considered not to contain any contaminant at a level that might have affected the purpose or integrity of the study.
Details on mating procedure:
Animals were paired on a 1 male: 1 female basis within each dose group, for a period of up to fourteen days. Cage tray-liners were checked each morning for the presence of ejected copulation plugs and each female was examined for the presence of a copulation plug in the vagina. A vaginal smear was prepared for each female and the stage of estrus or the presence of sperm was recorded. The presence of sperm within the vaginal smear and/or vaginal plug in situ was taken as positive evidence of mating (Day 0 of gestation) and the males were subsequently returned to their original holding cages. Mated females were housed individually during the period of gestation and lactation.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
HPLC/UV- single peak
System: Agilent tech 1200, autosampler and workstation
Column: Eclispse XDB C18 (150 x 4.6 mm id)
Mobile phase: ACN:H2O (70:30) v/v
Fow rate: 1mL/min
Wavelength: 220 nm
Injection volume: 25µL
Retention time: ~ 4.7 mins
Duration of treatment / exposure:
approximately six weeks (males) and up to eight weeks (females) (including a two week pre-pairing phase, pairing, gestation and early lactation for females) at dose levels of 5, 15 and 25 mg/kg bw/day. A control group was dosed with vehicle alone (Polyethylene glycol 400).
Frequency of treatment:
daily
Details on study schedule:
- F1 parental animals not mated until 2 weeks after selected from the F1 litters.
- Age at mating of the mated animals in the study: 13 weeks males , 16 weeks females
Doses / concentrationsopen allclose all
Dose / conc.:
0 mg/kg bw/day (nominal)
Dose / conc.:
5 mg/kg bw/day (nominal)
Dose / conc.:
15 mg/kg bw/day (nominal)
Dose / conc.:
25 mg/kg bw/day (nominal)
No. of animals per sex per dose:
12
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: from preliminary study, and after discussing with sponsor
- Rationale for animal assignment (if not random): random
Positive control:
none

Examinations

Parental animals: Observations and examinations:
DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule:
All animals were examined for overt signs of toxicity, ill-health and behavioral change immediately before dosing, soon after dosing, and one hour after dosing during the working week and at weekends (except for females during parturition where applicable). All observations were recorded.

BODY WEIGHT: Yes
- Time schedule for examinations:
Individual body weights were recorded on Day 1 (prior to dosing) and then weekly for males until termination and weekly for females until pairing. During pairing phase females were weighed daily until mating was confirmed. Body weights were then recorded for females on Days 0, 7, 14 and 20 post coitum, and on Days 1, 4, 7 and 14 post partum. Body weights were also recorded at terminal kill.

FOOD CONSUMPTION AND COMPOUND INTAKE:
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: Yes

FOOD EFFICIENCY:
- Body weight gain in kg/food consumption in kg per unit time X 100 calculated as time-weighted averages from the consumption and body weight gain data: Yes, During the pre-pairing period, weekly food consumption was recorded for each cage of adults. This was continued for males after the mating phase. For females showing evidence of mating, food consumption was recorded for the periods covering post coitum Days 0-7, 7-14 and 14-20. For females with live litters, food consumption was recorded for the periods covering post partum Days 1-4, 4-7 and 7-14. Food efficiency (the ratio of body weight change/dietary intake) was calculated retrospectively for males throughout the study period (with the exception of the mating
phase) and for females during the pre-pairing phase. Due to offspring growth and milk production, food efficiency could not be accurately calculated during gestation and lactation.

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): Yes
- Time schedule for examinations:Water intake was measured daily during the pre-pairing phase of the study. Intergroup differences did not indicate any need for more formal gravimetric measurements.


OPHTHALMOSCOPIC EXAMINATION:No

HAEMATOLOGY: Yes The following parameters were measured on blood collected into tubes containing potassium EDTA anti-coagulant:
Hemoglobin (Hb)
Erythrocyte count (RBC)
Hematocrit (Hct)
Erythrocyte indices - mean corpuscular hemoglobin (MCH)
- mean corpuscular volume (MCV)
- mean corpuscular hemoglobin concentration (MCHC)
Total leukocyte count (WBC)
Differential leukocyte count - neutrophils (Neut)
- lymphocytes (Lymph)
- monocytes (Mono)
- eosinophils (Eos)
- basophils (Bas)
Platelet count (PLT)
Reticulocyte count (Retic)
Prothrombin time (CT) was assessed by ‘Innovin’ and Activated partial thromboplastin time (APTT) was assessed by ‘Actin FS’ using samples collected into sodium citrate solution (0.11 mol/L).

CLINICAL CHEMISTRY: Yes.
The following parameters were measured on plasma from blood collected into tubes containing lithium heparin anti-coagulant:
Urea Inorganic phosphorus (P)
Glucose Aspartate aminotransferase (ASAT)
Total protein (Tot.Prot.) Alanine aminotransferase (ALAT)
Albumin Alkaline phosphatase (AP)
Albumin/Globulin (A/G) ratio (by calculation) Creatinine (Creat)
Sodium (Na+) Total cholesterol (Chol)
Potassium (K+) Total bilirubin (Bili)
Chloride (Cl-) Bile acids
Calcium (Ca++)


URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: Yes
- Time schedule for examinations:
Prior to the start of treatment and at weekly intervals thereafter, all animals were observed for signs of functional/behavioral toxicity. These observations were performed on mated females on Days 4, 11 and 18 post coitum and for littering females on Days 4 and 12 post partum. Functional performance tests were also performed on five selected males and females from each dose level, prior to termination, together with an assessment of sensory reactivity to various stimuli. Detailed individual clinical observations were performed for each animal using a purpose built arena. The following parameters were observed:
Gait Hyper/Hypothermia
Tremors Skin color
Twitches Respiration
Convulsions Palpebral closure
Bizarre/Abnormal/Stereotypic behavior Urination
Salivation Defecation
Pilo-erection Transfer arousal
Exophthalmia Tail elevation
Lachrymation
This test was developed from the methods used by Irwin (1968) and Moser et al (1988). The scoring system used is outlined in The Key to Scoring System and Explanation for Behavioral Assessments and Sensory Reactivity Tests.
Oestrous cyclicity (parental animals):
Vaginal smears were taken daily for females throughout the two week pre-pairing treatment period and in the morning of the day of necropsy. The stage of the estrous cycle was recorded for each day.
Sperm parameters (parental animals):
not specified
Litter observations:
On completion of parturition (Day 0 post partum), the number of live and dead offspring was recorded. Offspring were individually identified within each litter by tattoo on Day 1 post partum.
For each litter the following was recorded:
i. Number of offspring born
ii. Number of offspring alive recorded daily and reported on Days 1, 4, 7 and 13 post partum
iii. Sex of offspring on Days 1, 4 and 13 post partum
iv. Clinical condition of offspring from birth to Day 13 post partum
v. Individual offspring weights on Days 1, 4, 7 and 13 post partum (litter weights were calculated retrospectively from this data)
Postmortem examinations (parental animals):
Adult males were killed by intravenous overdose of suitable barbiturate agent followed by exsanguination on Day 44 or 45. Adult females were killed by intravenous overdose of suitable barbiturate agent followed by exsanguination on Day 14 post partum. Surviving offspring were terminated by carbon dioxide asphyxiation followed by cervical dislocation on Day 13 post partum. Offspring required for blood sampling were terminated by cervical dislocation with death confirmed by decapitation during the sampling procedure with blood samples collected immediately following decapitation.
For all females, the uterus was examined for signs of implantation and the number of uterine implantations in each horn was recorded. This procedure was enhanced; as necessary, by staining the uteri with a 0.5% ammonium polysulphide solution (Salewski 1964).
All adult animals and offspring, including those dying during the study, were subjected to a full external and internal examination, and any macroscopic abnormalities were recorded. Examination of offspring was restricted to a macroscopic external examination except where abnormalities were observed, then an additional internal examination was performed.
Postmortem examinations (offspring):
All offspring were examined externally; where external observations were detected an internal necropsy was performed.
Statistics:
Where considered appropriate, quantitative data was subjected to statistical analysis to detect the significance of intergroup differences from control; statistical significance was achieved at a level of p<0.05. Statistical analysis was performed on the following parameters:
Grip Strength, Motor Activity, Body Weight, Body Weight Change, Food Consumption during gestation and lactation, Pre-Coital Interval, Gestation Length, Litter Size, Litter Weight, Sex Ratio, Corpora Lutea, Implantation Sites, Implantation Losses, Viability Indices, Offspring Body Weight, Offspring Body Weight Change, Offspring Developmental Parameters, Hematology, Blood Chemistry, Absolute Organ Weights, Body Weight-Relative Organ Weights and Thyroid Hormone (Thyroxine).
Data were analyzed using the decision tree from the ProvantisTM Tables and Statistics Module
Reproductive indices:
i. Pre-coital Interval
ii. Fertility Indices
i. Gestation Length
ii. Parturition Index
Offspring viability indices:
i. Implantation Losses (%)
ii. Live Birth and Viability Indices
iii. Sex Ratio (% males)

Results and discussion

Results: P0 (first parental generation)

General toxicity (P0)

Clinical signs:
no effects observed
Description (incidence and severity):
There were considered to be no clinical signs which could be directly related to systemic toxicity of the test item in animals of either sex treated with 5, 15 or 25 mg/kg bw/day.
Animals of either sex treated with 25 mg/kg bw/day showed occasional instances of increased salivation (four males and nine females) from Day 9 to Day 41 (males) and Day 6 to Day 56 (females). Occasional instances of noisy respiration (ten males and all females) were also noted in these animals from Day 4 to Day 42 (males) and in female animals from Day 2 until Day 40. Two male animals exhibited staining around the snout on Day 23.
One male and one female treated with 15 mg/kg bw/day exhibited increased salivation for one day only. Isolated instances of noisy respiration was noted in six males and six female animals from Day 4 to Day 33 (males) and from Day 2 to Day 45 (females).
One female treated with 5 mg/kg bw/day exhibited noisy respiration for one day only. Generalized fur loss was also noted in one female animal from Day 32 to Day 53, as this was only noted in one female animal it was considered not to be related to treatment with the test item. No such effects were noted in the respective male animals from this treatment group.
One control male and one control female animal exhibited noisy respiration on one or two days respectively.
Increased salivation is commonly observed following the oral administration of an unpalatable or slightly irritant test item formulation and is generally considered to be of no toxicological importance. Noisy respiration was considered to be a possible consequence of the increased salivation or possible difficulty in dosing these animals rather than any underlying toxicological effect of the test item.
No clinical signs were apparent in any of the control animals prior to being found dead.
Dermal irritation (if dermal study):
not examined
Mortality:
mortality observed, non-treatment-related
Description (incidence):
Two control female animals were found dead on Days 12 or 18.
There were no further unscheduled deaths during this study.
Body weight and weight changes:
no effects observed
Description (incidence and severity):
Group mean weekly body weights and standard deviations are given in Table 7 and are presented graphically in Figure 1 and Figure 2. Group mean weekly body weight gains and standard deviations are given in Table 8 (statistically significant differences are indicated). Individual data are given in Appendix 7 and Appendix 8.
There was considered to be no adverse effect of treatment on body weight development in treated male animals throughout the study.
Males treated with 25 mg/kg bw/day showed a statistically significant (p<0.05) increase in body weight gain during the third week of treatment. An increase in body weight gain is considered not to be an adverse effect of treatment and therefore is considered not to be toxicologically significant.
Male animals treated with 15 mg/kg bw/day exhibited reductions in body weight gain during the first two weeks of treatment but without achieving statistical significance. As no such effects were noted in animals treated with 25 mg/kg bw/day this finding is considered to be most likely due to normal biological variation rather than any underlying systemic toxicity elicited by the test item.
There were considered to be no effects on body weight development in treated females during maturation. There were also considered to be no adverse effects on body weight gain during the gestation period for any of the treated female animals.
During the lactation phase of the study females treated with 25 mg/kg bw/day exhibited reductions in body weight gains (without achieving statistical significance) when compared to control animals from Day 1 to Day 4 and from Days 4 to 7. Body weight gains were then comparable to control in these animals during the second week of lactation, however, overall body weight gain was still 22% lower than control.
Females treated with 25 mg/kg bw/day exhibited slight reductions (without achieving statistical significance) in body weight gain throughout gestation when compared to control which resulted in a 13% reduction in overall body weight gain. This may be a reflection of slightly lower litter sizes in this treatment group when compared to control. With two females removed from group means which exhibited much lower litter size (and where considered atypical for the group as a whole) overall body weight gain was only 7% lower than control.
Females treated with 5 and 15 mg/kg bw/day exhibited slight reductions (without achieving statistical significance) in body weight gain during the first week of gestation, however, a dose relationship was not apparent. Body weight gains were then comparable to control during the next two weeks of treatment. Overall body weight gain for these animals were 6% and 5% lower than control for animals treated with 5 and 15 mg/kg bw/day respectively.
There was considered to be no adverse effect of treatment on body weight development during lactation in females treated with 5 or 15 mg/kg bw/day. Females treated with 15 mg/kg bw/day exhibited a reduction in body weight gain during the second week of lactation and females treated with 5 mg/kg bw/day exhibited a reduction in body weight gain from Days 4 to 7 of lactation (statistical significance was not achieved in either instance), however, dose relationships were not apparent and overall body weight gain remained generally comparable to control.
Food consumption and compound intake (if feeding study):
no effects observed
Description (incidence and severity):
There were no effects in food consumption in any treated male throughout the treatment period or in any treated female animals during maturation or gestation.
Females treated with 25 mg/kg bw/day exhibited reductions in food consumption during lactation which achieved statistical significance (p<0.01) from Days 4 to 7.
There were considered to be no effects noted in food consumption during lactation for females treated with 5 or 15 mg/kg bw/day.
Food efficiency:
no effects observed
Description (incidence and severity):
There was considered to be no effects in food conversion efficiency in any males or in any treated female animal during the pre-pairing phase of the study.
Water consumption and compound intake (if drinking water study):
no effects observed
Description (incidence and severity):
There were considered to be no effects in water consumption for any treated animal.
Ophthalmological findings:
not examined
Haematological findings:
no effects observed
Description (incidence and severity):
There were considered to be no adverse effects of treatment on the hematological parameters measured.
Statistically significant increases in erythrocytes (p<0.05) were noted in males treated with 15 and 25 mg/kg bw/day and statistically significant increases (p<0.05) in clotting time was noted in male animals treated with 5 and 15 mg/kg bw/day. Animals of either sex from all treatment groups exhibited statistically significant increases in reticulocytes (p<0.01). Statistically significant reductions in leukocytes and neutrophils (p<0.05) were also noted in females from all treatment groups and from the 25 mg/kg bw/day treatment group respectively.
With the exception of reticulocyte counts which appeared to be dose related, no dose relationship was apparent in any of these findings and all values were found to be within the historical control data ranges. As there were also no histopathological correlates these findings are considered not to be of any toxicological significance.
Clinical biochemistry findings:
no effects observed
Description (incidence and severity):
There were considered to be no adverse effects of treatment on the blood chemical parameters measured.
Males from all treatment groups exhibited statistically significant increases in chloride concentration (p<0.01) and alkaline phosphatase (p<0.05). Whilst males treated with 5 mg/kg bw/day exhibited statistically significant increases in albumin/globulin ratio. None of these findings exhibited a dose relationship and with the exception of two control animals which exhibited extremely low alkaline phosphatase values, all values were within the historical control data range. Male animals treated with 25 mg/kg bw/day exhibited statistically significant (p<0.05) reductions in urea, however, all values were within the historical control data ranges. As there were also no histopathological correlates these findings are considered not to be of any toxicological significance.
No such findings were noted in the female animals from any treatment group.
Urinalysis findings:
not examined
Behaviour (functional findings):
no effects observed
Description (incidence and severity):
There were considered to be no toxicologically significant effects in the behavioral parameters measured. A small number of instances of noisy respiration were noted during the behavioral assessments in control males and in animals of either sex treated with 5 and 25 mg/kg bw/day. One control animal also exhibited an isolated occurrence of diarrhoea.
There were no changes in the behavioral parameters in animals of either sex treated with 15 mg/kg bw/day.
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
no effects observed
Description (incidence and severity):
There were considered to be no treatment related changes in functional performance considered to be related to treatment at 5, 15 or 25 mg/kg bw/day.
Male animals treated with 25 mg/kg bw/day exhibited a statistically significant increase (p<0.05) in hind limb grip strength and a statistically significant increase (p<0.01) in the last 20% of activity monitoring. As the increase in hind limb grip strength was only noted in one out of the three runs completed and there were no clinical signs observed or any histopathological correlates to signify a neurotoxic effect of the test item these two incidences were considered to be incidental and of no toxicological relevance.
Histopathological findings: non-neoplastic:
no effects observed
Histopathological findings: neoplastic:
not examined
Description (incidence and severity):
A complete histopathology phase report is presented in Annex 1.
There were no adverse effects of treatment noted at histopathological examination.
Mesenteric Lymph node
Multifocal histiocytosis (histiocyte aggregates) were present in 1/5 males treated with 15 or 25 mg/kg bw/day and 2/5 females treated with 15 mg/kg bw/day and 3/5 females treated with 25 mg/kg bw/day. This was a minimal change and is known to occur in response to the oral administration of some test items. Without further evidence of pathological change, necrosis or abscess formation, it is considered not to affect the functionality of the lymph node and generally considered to be non-adverse.
Uterus
Pigmentation at the involuting implantation sites was variable but after examination from all females no consistent differences were found and changes were therefore considered to be incidental.
Other effects:
not examined

Reproductive function / performance (P0)

Reproductive function: oestrous cycle:
no effects observed
Description (incidence and severity):
There was no effect of treatment with the test item at any dose level on the nature of estrous cycle. All females showed regular cycles over the pre-pairing phase of the study. There were also no intergroup differences in the stage of estrus on the day of necropsy.
Reproductive function: sperm measures:
not examined
Reproductive performance:
no effects observed
Description (incidence and severity):
No treatment-related effects were detected in mating performance.

Effect levels (P0)

Key result
Dose descriptor:
NOAEL
Effect level:
25 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
gross pathology
histopathology: non-neoplastic
reproductive function (oestrous cycle)
reproductive performance

Target system / organ toxicity (P0)

Key result
Critical effects observed:
no
System:
other: various
Organ:
other:

Results: F1 generation

General toxicity (F1)

Clinical signs:
no effects observed
Description (incidence and severity):
The clinical signs apparent for offspring on the study were typical for the age observed. Neither the incidence nor distribution of these observations indicated any effect of maternal treatment on offspring development in any treatment group.
Dermal irritation (if dermal study):
not examined
Mortality / viability:
no mortality observed
Body weight and weight changes:
no effects observed
Description (incidence and severity):
There was considered to be no adverse effect of treatment with the test item indicated by offspring body weight, body weight gain or litter weights.
As a consequence of the slightly reduced litter size from animals treated with 25 mg/kg bw/day, litter weights on Days 1, 4, 7, and 13 post partum were reduced when compared to controls but without achieving statistical significance. These findings were considered to be slightly overemphasized by the two animals which only gave birth to four or five offspring. When these two animals were removed, group mean litter weights were comparable to control.
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:
no effects observed
Description (incidence and severity):
There were considered to be no adverse effects of treatment on the hematological parameters measured.
Clinical biochemistry findings:
no effects observed
Urinalysis findings:
not examined
Sexual maturation:
no effects observed
Description (incidence and severity):
There was no effect of treatment with the test item indicated by ano-genital distance on Day 1 post partum or visible nipple count in male offspring on Day 13 post partum from any treatment group.
Organ weight findings including organ / body weight ratios:
no effects observed
Histopathological findings:
no effects observed
Other effects:
not examined

Developmental neurotoxicity (F1)

Behaviour (functional findings):
no effects observed

Developmental immunotoxicity (F1)

Developmental immunotoxicity:
not examined

Effect levels (F1)

Key result
Dose descriptor:
NOAEL
Generation:
F1
Effect level:
25 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
viability
sexual maturation
clinical signs
mortality
body weight and weight gain
haematology
clinical biochemistry
histopathology: non-neoplastic

Target system / organ toxicity (F1)

Key result
Critical effects observed:
no

Overall reproductive toxicity

Key result
Reproductive effects observed:
no
Treatment related:
no

Applicant's summary and conclusion

Conclusions:
The ‘No Observed Adverse Effect Level’ (NOAEL) for reproductive toxicity was considered to be 25 mg/kg bw/day.