Registration Dossier

Diss Factsheets

Administrative data

Key value for chemical safety assessment

Effects on fertility

Description of key information

No significant adverse effects of the registration substance were revealed when tested in a reproductive toxicity screening tests according to OECD TG 421 in rats at doses up to 1000 mg/kg body weight. Based on the results tthe NOAEL (No Observed Adverse Effect Level) for reproduction/developmental toxicity was established at 1000 mg/kg/day for both males and females.

Link to relevant study records
Reference
Endpoint:
screening for reproductive / developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2012
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: guideline-conform study under GLP without deviations
Qualifier:
according to guideline
Guideline:
OECD Guideline 421 (Reproduction / Developmental Toxicity Screening Test)
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Limit test:
no
Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River, S.r.l. Calco, (Lecco), Italy.
- Age at study initiation: 8-9 wks
- Weight at study initiation: (P) Males: 255.13-257.18 g; Females: 193.39-193.90 g
- Fasting period before study: no
- Housing: 5 per cage (polysulfone solid bottomed cages)
- Diet (e.g. ad libitum): yes
- Water (e.g. ad libitum): yes
- Acclimation period: approximately 2 wks

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22°C +/- 3°C
- Humidity (%): 30-70%
- Air changes (per hr): approximately 15-20 changes
- Photoperiod (hrs dark / hrs light): 12 hrs

IN-LIFE DATES: 20 June - 12 August 2012
Route of administration:
oral: gavage
Vehicle:
water
Details on exposure:
The required amount of Hostapon TPHC was suspended in purified water at the final concentrations of 10, 30 and 100 mg/mL. The formulations
were prepared daily and the concentrations were calculated and expressed in terms of test item as supplied.
Details on mating procedure:
- M/F ratio per cage: one to one
- Length of cohabitation: up to 14 days
- Proof of pregnancy: vaginal plug / sperm in vaginal smear referred to as day 0 of pregnancy
- Further matings after two unsuccessful attempts: no
- After successful mating each pregnant female was caged (how): single
- Any other deviations from standard protocol: no
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Chemical analysis was carried out by the Analytical Chemistry Department at RTC according to a validated method (RTC Study No. 91450).
The validation of the formulation procedure for Hostapon TPHC was performed at 10, 30 and 100 mg/mL. The results were obtained in RTC Study
No. 91450 and all levels were within the acceptability limits for concentration and homogeneity.
The stability was found to be 24 hours at room temperature in the concentration range of 10 to 100 mg/mL in the same study.
Samples of the formulations prepared on Weeks 1 and 6 (when all females were present) of the present study were also analysed to check the concentration and homogeneity.
The overall results of the analyses were within the limits of acceptance stated in RTC’s SOPs for concentration (90-110%) and homogeneity (CV <10%).
The software used for this activity was Analyst 1.5.2 (AB Sciex).
Duration of treatment / exposure:
Males
Animals were dosed once a day, 7 days a week, for 2 consecutive weeks prior to pairing and thereafter through the day before necropsy.
Dose volumes were adjusted once per week for each animal according to the last recorded body weight.

Females
Animals were dosed once a day, 7 days a week, for 2 consecutive weeks prior to pairing, with the exception of female no. 91480067 which was dosed for 15 consecutive days, and thereafter during pairing, post coitum and post partum periods until Day 3 post partum. Dose volumes were adjusted once per week for each animal according to the last recorded body weight.
During the gestation period, dose volumes were calculated according to individual body weight on Days 0, 7, 14 and 20 post coitum and on Day 1 post partum. Thereafter individual dose volumes remained constant.
Frequency of treatment:
Once a day
Details on study schedule:
Males were treated for a total of 33 days including 2 weeks prior to pairing and continuously thereafter up to the day before necropsy.
Females were dosed throughout the study including 2 weeks before pairing, thereafter during pairing, gestation and lactation periods until Day 3
post partum.
Remarks:
Doses / Concentrations:
100, 300, 1000 mg/kg body weight
Basis:
nominal in water
No. of animals per sex per dose:
10 animals/sex/group
Control animals:
yes, concurrent vehicle
Details on study design:
Dose selection rationale: The dosages of 100, 300 and 1000 mg/kg/day were selected in consultation with the Sponsor based on preliminary dose-range-finder. The oral route was selected as it is a possible route of exposure of the test item in man.

Parental animals: Observations and examinations:
Mortality
Throughout the study, all animals were checked early in each working day and again in the afternoon. At weekends and Public Holidays a similar procedure was followed except that the final check was carried out at approximately mid-day.

Clinical signs
All clinical signs were recorded for individual animals.
Once before commencement of treatment and at least once daily during the study, each animal was observed and any clinical signs recorded. Observations were performed at the same time interval each day, the interval was selected taking into consideration the presence of post-dose reactions.

Body weight
Males were weighed weekly from allocation to termination.
Females were weighed weekly from allocation to positive identification of mating and on Days 0, 7, 14 and 20 post coitum. Dams were also weighed on Days 1 and 4 post partum.

Food consumption
Food consumption was recorded at weekly intervals by each cage of rats from allocation to pairing. For female animals, food consumption was also recorded on Days 7, 14 and 20 post coitum, starting from Day 0 post coitum and on Day 4 post partum (starting from Day 1 post partum).
Oestrous cyclicity (parental animals):
Vaginal smears were taken daily in the morning from the first day of treatment and up to the end of the mating period, until a positive identification of mating was made. The vaginal smear data were examined to determine the following:
a) anomalies of the oestrous cycle;
b) the pre-coital interval (i.e., the number of nights paired prior to the detection of mating).
Sperm parameters (parental animals):
Parameters examined in all control and high dose parental male generations:
Epididymides, testis, prostate gland and seminal vesicles weights, morphological evaluation of the seminiferous epithelium (staging of the
spermatogenic cycle)

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. Live pups were individually weighed on Days 1 and 4
post partum. Observation was performed once daily for all litters.

GROSS EXAMINATION OF DEAD PUPS:
yes, for external and internal abnormalities
Postmortem examinations (parental animals):
SACRIFICE
- Male animals: All surviving animals were killed after the mating of all females (i.e. after 33 days of treatment).
- Maternal animals: The females with live pups were killed on Day 4 post partum

GROSS NECROPSY
The clinical history of the males and females of the parental generation was studied and a detailed post mortem examination was conducted
(including examination of the external surface and orifices).
Changes were noted, the requisite organs weighed (excluding animals sacrificed for humane reasons or found dead) and the required tissue samples
preserved in fixative and processed for histopathological examination.

All females were examined also for the following:
a) number of visible implantation sites (pregnant animals);
b) number of corpora lutea (pregnant animals).

The uterus of female no. 91480063 with no visible implantations was immersed in a 20% solution of ammonium sulphide to reveal evidence of
implantation.

HISTOPATHOLOGY / ORGAN WEIGHTS
From all animals completing the scheduled test period the organs were dissected free of fat and weighed (Annex 1 of the study protocol). The ratios of organ weight to terminal body weight
were calculated for each animal.

Tissues fixed and preserved
Samples of all the tissues listed in the Annex 1 of the study protocol were fixed and preserved in 10% neutral buffered formalin (except testes and epididymides which were fixed in modified Davidson's fluid) from all animals.

Histopathological examination and staging of spermatogenic cycle
The tissues required for histopathological examination are listed in Annex 1 of the study protocol. After dehydration and embedding in paraffin wax,
sections of the tissues were cut at 5 micrometre thickness and stained with haematoxylin and eosin.
In addition, the testes and epididymides were cut at 2-3 micrometre thickness and stained with Periodic Acid Schiff (PAS). The morphological evaluation of the seminiferous epithelium (staging of spermatogenic cycle) was performed.
The examination was restricted as detailed below:

a) Tissues from all animals in the control and high dose groups killed at term.
b) Tissues from all animals killed or dying during the treatment period.
c) All abnormalities in all groups.



Postmortem examinations (offspring):
Pups that had completed the scheduled test period (Day 4 post partum) were euthanised by intraperitoneal injection of Thiopenthal.

All pups found dead in the cage were examined for external and internal abnormalities. Sex was also confirmed by gonadal inspection.
All live pups were killed and examined for external abnormalities and sex confirmation by gonadal inspection.
Statistics:
Standard deviations were calculated as appropriate. For continuous variables the significance of the differences amongst group means was assessed by Dunnett’s test or a modified t test, depending on the homogeneity of data.
The non-parametric Kruskal-Wallis analysis of variance was used for the other parameters.
The criterion for statistical significance was p<0.05.
Reproductive indices:
Males
Copulatory Index (%) =no. of animals mated/ no. of animals paired x 100

Fertility Index (%)=no. of males which induced pregnancy/no. of males paired x 100

Females
Copulatory Index (%) =no. of animals mated/no. of animals paired x 100

Fertility Index (%) =no. of pregnant females/no. of females paired x 100

Males and females
Copulatory Interval = Mean number of days between pairing and mating

Offspring viability indices:
Pre-implantation loss was calculated as a percentage from the formula:

(No. of corpora lutea - No. of implantations)/ No. of corpora lutea x 100

Pre-birth loss was calculated as a percentage from the formula:

(No. of visible implantations - total litter size at birth )/ No. of visible implantations x 100

Pup loss at birth was calculated as a percentage from the formula:

(Total litter size - live litter size)/ Total litter size x 100

Cumulative pup loss on Day 4 post partum was calculated as a percentage from the formula:

(Total litter size at birth - live litter size at Day 4)/Total litter size at birth x 100

Sex ratios were calculated at birth and on Day 4 post partum and were presented as the percentage of males per litter.
Clinical signs:
no effects observed
Mortality:
mortality observed, non-treatment-related
Description (incidence):
Two high dose animals, one male and one female, were found dead on Day 8 of the study and on Day 0 post coitum, respectively. The cause of death for these two animals was attributed to a mis-dosing.
Body weight and weight changes:
no effects observed
Food consumption and compound intake (if feeding study):
no effects observed
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
not examined
Clinical biochemistry findings:
not examined
Urinalysis findings:
not examined
Behaviour (functional findings):
no effects observed
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
no effects observed
Histopathological findings: non-neoplastic:
no effects observed
Histopathological findings: neoplastic:
not examined
Other effects:
not examined
Reproductive function: oestrous cycle:
no effects observed
Reproductive function: sperm measures:
no effects observed
Reproductive performance:
no effects observed
Mortality
Two high dose animals were found dead during the study: one male (no. 91480068) on Day 8 of the study and one female (no. 91480063) on Day 0 post coitum. The cause of death was attributed to a mis-dosing on the basis of the macroscopic and histopathological findings.
All females were found pregnant at necropsy with the exception of the found dead animal no. 91480063 (Group 4).
The number of females with live pups on Day 4 post partum was: 10 in each of the control, low and mid- dose groups and 9 in the high dose group.

Clinical signs and observation of cage tray

Salivation was observed in the majority of high dose animals of both sexes throughout the study and occasionally in the low and mid-dose groups. This sign appeared early after dosing.
In one occasion, difficulty in breathing (dyspnoea) was observed in two high dose males (animal nos. 91480078 and 91480080).
No other relevant signs were recorded.
In several occasions, soft faeces and red/brown staining in the cage tray were noted in treated groups during the mating phase.
These data were not tabulated but retained as study raw data.

Body weight and body weight gain
Body weight and body weight gain were comparable between the groups throughout the study for both males and females.

Food consumption
Measurement of food consumption did not reveal relevant differences between the groups.

Oestrus cycle, reproductive parameters, pairing combination and mating performance
Measurements of oestrus cycle, pre-coital intervals, copulatory and fertility indices did not show differences between treated and control groups.

Implantation, pre-birth loss data and gestation length of females
No significant differences were found in the number of implantations, corpora lutea, total litter size, pre-implantation and pre-birth loss between control and treated groups.
Gestation length was comparable between groups.

Terminal body weight and organ weights
No differences were found in terminal body weight nor in organ weights between the groups.

Macroscopic observations

Unscheduled deaths
A high dose male (no. 91480068) and female (no. 91480063) died during the experimental phase.
At post mortem examination, the most relevant changes detected in the male or female rat were: red or brown staining in the muzzle, incomplete collapse, red colour or multiple dark areas in the lungs or red fluid in the thoracic cavity or red colour in the cervical lymph nodes or in the thymus.
Such changes, also confirmed by histopathological examination, were considered related to a possible misdosing and as consequence the factor contributory to death.

Final sacrifice
No relevant changes were detected at post mortem examination in treated animals, when compared with controls.

Microscopic observations
No treatment-related changes were seen in selected organs/tissues evaluated in males or females receiving Hostapon TPHC, sacrificed at the end of
treatment period, nor in the abnormalities detected in all groups at post mortem examination.
The sporadic lesions reported in single treated animals were considered to be an expression of spontaneous and/or incidental pathology, commonly seen in this species and age under our experimental conditions.

Seminiferous tubules were evaluated with respect to their stage in the spermatogenic cycle and to the integrity of the various cell types within the different stages; regular layering in the germinal epithelium was noted.
Key result
Dose descriptor:
NOAEL
Effect level:
1 000 mg/kg bw/day
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: On the basis of the results obtained in the study, the NOAEL (No Observed Adverse Effect Level) for reproduction/developmental toxicity could be considered 1000 mg/kg/day for both males and females.
Key result
Critical effects observed:
no
Clinical signs:
no effects observed
Mortality / viability:
no mortality observed
Body weight and weight changes:
no effects observed
Food consumption and compound intake (if feeding study):
not examined
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
not examined
Clinical biochemistry findings:
not examined
Urinalysis findings:
not examined
Sexual maturation:
not examined
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
no effects observed
Histopathological findings:
not examined
Behaviour (functional findings):
not examined
Developmental immunotoxicity:
not examined
Litter data at birth, on Day 1 and on Day 4 post partum of females and sex ratio of pups
Mean litter and pup weights were comparable between groups on Days 1 and 4 post partum.
No differences were found in sex ratio.

Clinical signs of pups
No differences were noted in the clinical signs observed in pups between groups.

Necropsy findings in decedent pups and in pups sacrificed on Day 4 post partum
No milk in stomach and autolysed organs in the abdomen were observed at necropsy in the decedent pups of control and treated groups.
No abnormalities were found in pups sacrificed on Day 4 post partum. Only one high dose pup had no milk in stomach.
Key result
Dose descriptor:
NOAEL
Generation:
F1
Effect level:
1 000 mg/kg bw/day (nominal)
Based on:
test mat.
Remarks on result:
not determinable due to absence of adverse toxic effects
Key result
Critical effects observed:
no
Reproductive effects observed:
not specified
Conclusions:
On the basis of the results obtained in the study, the NOAEL (No Observed Adverse Effect Level) for reproduction/developmental toxicity
could be considered 1000 mg/kg/day for both males and females.
Executive summary:

The registration substance was investigated according to OECD TG 421. The purpose of the study was to evaluate the reproduction/developmental toxicity of the test item in Sprague Dawley rats up to Day 4 post partum. The test item was administered orally, by gavage, at the dosages of 100, 300 and 1000 mg/kg/day. The treatment schedule included 2 weeks before pairing, during pairing, post coitum and post partum periods up to Day 3 post partum. Animals were administered for approximately 5 and 7 weeks for males and females, respectively. The parental animals were monitored for daily clinical signs, body weight, food consumption, oestrous cycle and mating performance. The dams were allowed to give birth and rear their offspring until Day 4 post partum. Macroscopic observation and histopathological examination were also performed. In parental animals, no relevant findings were found in life phase or at post mortem evaluation. The animals of the high dose group showed salivation early after dosing as major clinical sign. Body weight, body weight gain and food consumption were unaffected by treatment. No treatment-related findings were found at macroscopic and microscopic examinations. No abnormalities were found at the evaluation of the spermatogenic cycle. Likewise, no differences were noted in the reproductive performance including gonadal function, mating behaviour, conception, development of conceptus and parturition. The females had comparable length of gestation and gave births. Litter and mean pup weights were also comparable between groups and no relevant findings were observed in pups during the lactation period nor at post mortem examination. On the basis of the results obtained in the study, the NOAEL (No Observed Adverse Effect Level) for reproduction/developmental toxicity could be considered 1000 mg/kg/day for both males and females.

Effect on fertility: via oral route
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
1 000 mg/kg bw/day
Study duration:
subacute
Species:
rat
Quality of whole database:
Based on findings in a guideline conform reproductive screening study according to OECD TG 421 with the registration substance, neither effects with relevance to parental / maternal toxicity nor with regard to reproductive / developmental toxicity were observed. The NOAEL was established at 1000 mg/kg body weight per day for all endpoints. Additionally, an OECD TG 414 developmental toxicity study is included in this dossier.
Effect on fertility: via inhalation route
Endpoint conclusion:
no study available
Quality of whole database:
The available data base is considered sufficient for hazard / risk characterization. There are no indications of adverse effects on reproductive organs or tissues from the available data.
Effect on fertility: via dermal route
Endpoint conclusion:
no study available
Quality of whole database:
The available data base is considered sufficient for hazard / risk characterization.
Additional information

The registration substance was investigated according to OECD TG 421 for reproduction/developmental toxicity in Sprague Dawley rats up to Day 4 post partum. The test item was administered orally, by gavage, at the dosages of 100, 300 and 1000 mg/kg/day. The treatment schedule included 2 weeks before pairing, during pairing, post coitum and post partum periods up to Day 3 post partum. Animals were administered for approximately 5 and 7 weeks for males and females, respectively. The parental animals were monitored for daily clinical signs, body weight, food consumption, oestrous cycle and mating performance. The dams were allowed to give birth and rear their offspring until Day 4 post partum. Macroscopic observation and histopathological examination were also performed. In parental animals, no relevant findings were found in life phase or at post mortem evaluation. The animals of the high dose group showed salivation early after dosing as major clinical sign. Body weight, body weight gain and food consumption were unaffected by treatment. No treatment-related findings were found at macroscopic and microscopic examinations. No abnormalities were found at the evaluation of the spermatogenic cycle. Likewise, no differences were noted in the reproductive performance including gonadal function, mating behaviour, conception, development of conceptus and parturition. The females had comparable length of gestation and gave births. Litter and mean pup weights were also comparable between groups and no relevant findings were observed in pups during the lactation period nor at post mortem examination. On the basis of the results obtained in the study, the NOAEL (No Observed Adverse Effect Level) for reproduction/developmental toxicity could be considered 1000 mg/kg/day for both males and females.


Short description of key information:
The registration substance was tested in a guideline and GLP conform reproductive toxicity screening study according to OECD 421. No significant unspecific toxicity and mortality and no toxicologically relevant findings were noted for reproductive and developmental parameters in any dose group. The NOAEL for reproductive / developmental toxicity was considered to be 1000 mg/kg body weight. Additionally, a test proposal for an OECD TG 414 developmental toxicity study is included in this dossier.

Justification for selection of Effect on fertility via oral route:
Guideline study according to GLP. No derivations and/or confounders. Klimisch rating 1 representing reliability without restrictions. Information is valid and meet data requirements.

Effects on developmental toxicity

Description of key information
In an OECD prenatal developmental toxicity study, the No Observed Adverse Effect Level (NOAEL) of test item for maternal and embryofoetal toxicity in Wistar female rats via oral route was found to be the highest dose level employed i.e., 1000 mg/kg body weight/day
In the available GLP conform OECD TG 421 reproductive/developmental toxicity screening study, a NOAEL of 1000 mg/kg body weight per day was derived for male and female rats.
Link to relevant study records
Reference
Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2015-2016
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Limit test:
no
Species:
rat
Strain:
Wistar
Details on test animals or test system and environmental conditions:
The animal room was air-conditioned with adequate (above 10) air changes per hour The experimental room was continuously monitored for tem¬pera¬ture and relative humidity. The ranges for room tem¬pera¬ture and relative humidity were 20.5°C to 22.3°C and 51 to 65%, respectively. The animals were provided with a light cycle of 12 hours light and 12 hours dark.
Route of administration:
oral: gavage
Vehicle:
water
Details on exposure:
Dosing per gavage commenced on day 5 of gestation and continued until day 19 of gestation. Dose volume was 10 mL per kg body weight.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Stability and homogeneity analysis. Dose formulation anaylsis.
Details on mating procedure:
Animals were paired as one male and one female in the evening hours and following morning each female was examined for vaginal smear or the presence of a copulation plug in the vagina. The presence of sperm in the vaginal smear was taken as positive evidence of mating (Day 0 of gestation).The maximum mating period observed was of eight days. During the mating phase, animals were housed on one male: one female basis. After successful mating, the females were returned to their original cage and housed individually during gestation.
Duration of treatment / exposure:
Day 5 of gestation until day 19 of gestation
Frequency of treatment:
Once daily
Duration of test:
The dosing commenced on day 5 of gestation and continued until day 19 of gestation.
Dose / conc.:
0 mg/kg bw/day (nominal)
Dose / conc.:
100 mg/kg bw/day (nominal)
Dose / conc.:
300 mg/kg bw/day (nominal)
Dose / conc.:
1 000 mg/kg bw/day (nominal)
No. of animals per sex per dose:
24 female animals per dose group
Control animals:
yes, concurrent vehicle
Maternal examinations:
Clinical signs were recorded at same time(s) each day taking into consideration the peak period of anticipated effects after dosing. The condition of the animals was recorded including mortality, moribundity, pertinent behavioral changes, and all signs of overt toxicity. Body weights were examined during the acclimatization period once weekly, on first day of pairing and weekly thereafter as well as on gestation days 0, 3, 5, 8, 11, 14 17 and 20.
Ovaries and uterine content:
At termination, the uteri were removed and the pregnancy status of the animals was ascertained. Gravid uteri including the cervix were weighed. The uteri of females were examined for the presence and number of implantation sites and the number of corpora lutea in the ovaries were determined for pregnant animals. The uterine content was examined for number of resorption, live and/or dead foetuses. The degree of resorption was described in order to estimate the relative time of death of the conceptus. Uteri that appear non-gravid were further examined by 5% ammonium sulphide staining to reveal any early resorption or post implantation loss.
Fetal examinations:
Each foetus was subjected to external examination, which included all visible structures, surfaces and orifices (including the oral cavity). One-half of the fetuses (alternating foetuses within the litter) independent of sex were processed for skeletal alterations (odd number) and remaining half (even number) of each litter were examined for visceral (soft tissue) alterations. On completion of external examination the foetuses selected for skeletal evaluation were eviscerated and placed in 70% isopropyl alcohol.
Statistics:
Statistical analysis was performed using statplus program. All the data was checked for Normality with Shapiro-Wilk W test and for Homogeneity with Bartlett Chi-Square test. Each group of animals was subjected to Analysis of Variance (ANOVA). Values were given as mean ± standard deviation (SD).
Indices:
Pre-implantation loss, post-implantation loss, sex ratio, variation incidence
Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
In high dose (1000 mg/kg) group animals, burrowing followed by dullness and dyspnoea was observed shortly after dosing which subsided to normal after 3 hours post dosing from first day to last day of dosing. Only dullness was observed in some of the animals in intermediate dose (300 mg/kg) group following each gavage treatment which subsides to normal after 3 hours post dosing from first day to last day of dosing.
Mortality:
no mortality observed
Description (incidence):
No mortality was observed in any of the treated animal through out the scheduled treatment period.
Body weight and weight changes:
no effects observed
Description (incidence and severity):
No significant difference in the body weight and body weight gain (%) was observed in pregnant female animals of low (100 mg/kg), intermediate (300 mg/kg) and high dose (1000 mg/kg) groups. A statistically significant decrease in body weight was observed in high dose (1000 mg/kg) group when compared with low dose (100 mg/kg) group only on 20th day of gestation. However, this change was within the normal range with respect to age and sex of the strain of animals used. Hence, the changes in body weight and body weight gain were considered as biological variation without any toxicological relevance.
Detailed data see the tables in section "Any other information on results inc. tables"
Food consumption and compound intake (if feeding study):
no effects observed
Description (incidence and severity):
Feed consumption of pregnant female animals at low (100 mg/kg), intermediate (300 mg/kg) and high dose (1000 mg/kg) groups was not significantly different throughout the treatment period. A statistically significant decrease in feed consumption was observed in high dose (1000 mg/kg) group when compared with control (0 mg/kg), low (100 mg/kg) and intermediate dose (300 mg/kg) groups only on day 11th of gestation, and in control (0 mg/kg) group animals only on day 14th of gestation. These changes were within the normal range with respect to age and sex of the strain of animals used. Hence, these changes in feed consumption were considered as biological variation and thus not of toxicological relevance.
Detailed data see the tables in section "Any other information on results inc. tables"
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
not examined
Clinical biochemistry findings:
not examined
Urinalysis findings:
not examined
Behaviour (functional findings):
no effects observed
Description (incidence and severity):
Daily observations revealed no indications of general and functional pertinent behavioral changes.
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
no effects observed
Description (incidence and severity):
No abnormality was observed in any of the female animals from control, low, intermediate and high dose groups. Bursal cyst was observed in right ovary of one female animal of low dose group as an incidental finding.
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
not examined
Histopathological findings: neoplastic:
not examined
Other effects:
no effects observed
Number of abortions:
no effects observed
Description (incidence and severity):
No abortion noted througout the study period.
Data see tables in section "Any other information on results inc. tables"
Pre- and post-implantation loss:
no effects observed
Description (incidence and severity):
No pre- and/or post-implantation losses observed.
Data see tables in section "Any other information on results inc. tables"
Total litter losses by resorption:
no effects observed
Description (incidence and severity):
No litter losses by resorption observed.
Data see tables in section "Any other information on results inc. tables"
Early or late resorptions:
no effects observed
Description (incidence and severity):
No late resorptions observed.
Data see tables in section "Any other information on results inc. tables"
Dead fetuses:
no effects observed
Description (incidence and severity):
No dead fetuses observed.
Data see tables in section "Any other information on results inc. tables"
Changes in pregnancy duration:
no effects observed
Description (incidence and severity):
No changes on pregnancy duration observed.
Migrated Data from removed field(s)
Field "Effects on pregnancy duration" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsMaternalAnimals.MaternalDevelopmentalToxicity.EffectsOnPregnancyDuration): no effects observed
Field "Description (incidence and severity)" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsMaternalAnimals.MaternalDevelopmentalToxicity.DescriptionIncidenceAndSeverityEffectsOnPregnancyDuration): No effects on pregnancy duration observed.
Changes in number of pregnant:
no effects observed
Description (incidence and severity):
All the female animals of control, low, intermediate and high dose groups showed positive evidence of mating. At necropsy, three females each of control, high and low dose groups and four females of intermediate dose group were found non pregnant. As all dose groups are comparably involved and no dose response occurred, this finding is not of toxicological relevance.
Detailed data see tables in section "Any other information on results inc. tables"
Other effects:
no effects observed
Description (incidence and severity):
No significant difference was observed in the sex ratio of foetuses in low, intermediate and high dose groups or other reproductive indices.
Detailed data see tables in section "Any other information on results inc. tables"
Details on maternal toxic effects:
No maternal effects of toxicological relevance observed throughout the study period.
Key result
Dose descriptor:
NOAEL
Effect level:
1 000 mg/kg bw/day (nominal)
Based on:
test mat.
Basis for effect level:
other: overall findings
Remarks on result:
not determinable due to absence of adverse toxic effects
Key result
Abnormalities:
no effects observed
Fetal body weight changes:
no effects observed
Description (incidence and severity):
No significant difference was observed in litter and foetus weight of low, intermediate and high dose groups.
Detailed data see tables in section "Any other information on results inc. tables"
Migrated Data from removed field(s)
Field "Fetal/pup body weight changes" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsFetuses.FetalPupBodyWeightChanges): no effects observed
Field "Description (incidence and severity)" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsFetuses.DescriptionIncidenceAndSeverityFetalPupBodyWeightChanges): No changes in fetal/pub body weights recorded.
Detailed data see tables in section "Any other information on results inc. tables"
Reduction in number of live offspring:
no effects observed
Description (incidence and severity):
No significant differences were observed in early and late resorptions and in the number of live foetuses in all treatment groups.
Detailed data see tables in section "Any other information on results inc. tables"
Changes in sex ratio:
no effects observed
Description (incidence and severity):
No significant difference was observed in the sex ratio of foetuses in low, intermediate and high dose groups.
Detailed data see tables in section "Any other information on results inc. tables"
Changes in litter size and weights:
no effects observed
Description (incidence and severity):
No significant difference was observed in litter size and the foetus weights in low, intermediate and high dose groups.
Detailed data see tables in section "Any other information on results inc. tables"
Changes in postnatal survival:
not examined
External malformations:
no effects observed
Description (incidence and severity):
External examination of foetuses revealed one small sized foetus in control, six in low dose, eight in intermediate dose and three in high dose group. However, statistical significant differences in respective body weights were not observed. As the external variations observed were randomly distributed across the groups and in the absence of any dose response relationship, these findings were considered as incidental and of no toxicological relevance.
Detailed data see tables in section "Any other information on results inc. tables"
Skeletal malformations:
no effects observed
Description (incidence and severity):
Skeletal examination of foetuses revealed variations common in type and distribution for the strain of rats used. They were isolated and when expressed on a foetus/litter basis, no statistical significant differences between treatment groups and control animals were observed.
Skeletal examination at the dose levels of 100, 300 and 1000 mg/kg body weight thus did not indicate any test item-related effects.
Detailed data see tables in section "Any other information on results inc. tables"
Visceral malformations:
no effects observed
Description (incidence and severity):
Variations observed during visceral examination of foetuses revealed variations common in type and incidence for the strain of rats used and were randomly distributed across all groups. A dose response relationship was not observed. Therefore these findings were considered as incidental findings and of no toxicological relevance.
Detailed data see tables in section "Any other information on results inc. tables"
Other effects:
no effects observed
Description (incidence and severity):
No effects observed
Details on embryotoxic / teratogenic effects:
No effects observed
Key result
Dose descriptor:
NOAEL
Effect level:
1 000 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: overall effects
Remarks on result:
not determinable due to absence of adverse toxic effects
Key result
Abnormalities:
no effects observed
Key result
Developmental effects observed:
no

SUMMARY OF MORTALITY AND CLINICAL SIGNS

 

Group

1

2

3

4

Dose (mg/kg bw)

0

100

300

1000

Mortality

Nil

Nil

Nil

Nil

Clinical signs

Nil

Nil

Dullness

Excessive burrowing,

Dullness,

Dyspnoea

 

BODY WEIGHTS (G) – SUMMARY

FEMALE

Group

Body weights (G)

Gestation Day

 

0

3

5

8

11

14

17

20

Group 1

Mean

227.25

232.75

238.29

246.21

256.15

266.79

281.79

306.38

SD

10.01

9.73

9.59

9.57

11.22

12.63

18.22

25.60

Min

210.2

214.4

218.2

224.8

232.2

245.2

251.4

263.1

Max

244.2

250.2

253.8

260.4

270.2

290.2

323.4

352.1

N

21

21

21

21

21

21

21

21

Group 2

Mean

228.79

235.81

242.44

250.46

259.14

270.67

286.48

310.64

SD

10.46

10.67

11.56

11.34

12.33

15.51

15.11

17.30

Min

212.4

218.3

224.1

232.1

238.2

243.4

260.2

274.3

Max

244.6

251.4

267.1

273.8

282.2

294.1

310.5

338.2

N

21

21

21

21

21

21

21

21

Group 3

Mean

225.28

231.71

238.18

246.19

256.07

266.52

279.71

298.19

SD

11.39

10.20

10.37

11.19

11.86

13.16

14.74

19.09

Min

209.1

218.1

224.0

230.4

238.2

247.1

259.1

269.2

Max

245.3

249.2

256.8

270.4

281.3

292.1

310.1

332.1

N

20

20

20

20

20

20

20

20

Group 4

Mean

224.95

231.49

238.42

246.10

255.90

266.32

278.81

293.96#

SD

10.61

10.47

10.32

9.68

10.54

11.75

13.28

14.84

Min

210.6

220.1

226.3

233.2

240.1

249.2

259.2

268.1

Max

246.8

252.8

255.2

260.2

271.1

286.2

304.1

321.4

N

21

21

21

21

21

21

21

21

#Significant at p ≤ 0.05 level with group 2

BODY WEIGHT GAIN (%) - SUMMARY

FEMALE

Group

Body weights Gain (%)

Gestation Day

 

3

5

8

11

14

17

20

Group 1

Mean

2.43

4.88

8.39

12.77

17.44

24.02

34.91

SD

1.00

1.61

2.44

3.48

3.90

6.33

10.90

N

21

21

21

21

21

21

21

Group 2

Mean

3.08

5.99

9.52

13.30

18.32

25.28

35.84

SD

1.43

2.74

3.27

3.15

4.59

5.24

6.22

N

21

21

21

21

21

21

21

Group 3

Mean

2.89

5.77

9.33

13.72

18.35

24.19

32.41

SD

1.26

1.65

2.48

2.98

3.00

3.38

6.34

N

20

20

20

20

20

20

20

Group 4

Mean

2.92

6.02

9.47

13.84

18.50

24.03

30.82

SD

1.21

1.92

2.82

3.75

4.95

5.01

6.90

N

21

21

21

21

21

21

21

 

FEED CONSUMPTION - SUMMARY

FEMALE

Group

FEED CONSUMPTION (G/ANIMAL/DAY)

Gestation Day

 

0 - 3

3 - 5

5 - 8

8 - 11

11 - 14

14 - 17

17 - 20

Group 1

Mean

17.40

19.26

20.26

21.99

23.99

24.16

23.85

SD

1.48

3.45

1.87

1.72

2.15

2.79

2.88

Group 2

Mean

18.06

19.15

19.62

21.91

23.09

23.43

22.91

SD

2.03

3.96

2.76

2.48

1.48

1.82

2.88

Group 3

Mean

17.59

19.97

19.47

22.04

23.23

23.55

23.38

SD

2.53

5.38

3.28

2.58

2.40

2.26

3.17

Group 4

Mean

17.04

17.93

18.71

19.10*#@

21.41*

22.31

22.10

SD

2.10

4.12

3.00

3.57

3.17

2.42

2.54

*Significant at p ≤ 0.05 level with group 1

#Significant at p ≤ 0.05 level with group 2

@Significant at p ≤ 0.05 level with group 3

PREGNANCY STATUS

GROUP

G1

G2

G3

G4

DOSE (mg/kg bw)

0

100

300

1000

FEMALE ANIMALS MATED

24

24

24

24

PREGNANT FEMALE ANIMALS

21

21

20

21

 

MACROSCOPIC FINDINGS- FEMALE

GROUP

G1

G2

G3

G4

DOSE (mg/kg bw)

0

100

300

1000

ANIMALS EXAMINED

24

24

24

24

ANIMALS WITHOUT ABNORMALITY

24

24

24

24

ANIMALS AFFECTED

0

0

0

0

MATERNAL DATA - SUMMARY

 

Parameter

Group

G1

G2

G3

G4

Dose (mg/kg bw)

0

100

300

1000

No. of Dams

21

21

20

21

Gravid Uterine Weight

Mean

43.9637

53.2278

45.9287

49.9408

SD

16.0935

10.1451

14.7209

18.3332

No. of CL

Mean

11.96

12.92

12.92

12.58

SD

2.73

1.77

2.34

2.80

No. of Implantation

Mean

8.13

9.92

8.38

9.13

SD

4.24

4.15

4.83

4.76

No. of Foetuses

Mean

8.38

10.14

9.00

9.71

SD

3.35

2.31

3.36

3.98

Early Resorption

Mean

0.79

0.88

0.88

0.54

SD

0.88

1.15

1.08

0.88

Late Resorption

Mean

0.00

0.17

0.00

0.08

SD

0.00

0.64

0.00

0.28

Pre implantation loss

Mean

3.83

3.00

4.54

3.46

SD

4.31

3.41

4.32

3.71

%

30.90

24.41

35.89

29.12

Post implantation loss

Mean

0.79

1.04

0.92

0.63

SD

0.88

1.20

1.10

1.01

%

12.59

14.49

12.01

9.14

Dam with resorption/s

Number

13

12

12

8

LITTER DATA - SUMMARY

 

Parameter

Group

G1

G2

G3

G4

Dose (mg/kg bw)

0

100

300

1000

No. of Dams

24

24

24

24

No. of Litters

21

21

20

21

Total No. of Foetuses

176

213

180

204

Mean Litter Size

8.38

10.14

9.00

9.71

No. of Live Foetuses

Number

176

213

179

204

Mean

8.38

10.14

8.95

9.71

SD

3.35

2.31

3.32

3.98

No. of Dead Foetuses

Number

0

0

1

0

Mean

0

0

0.05

0

SD

0.00

0.00

0.22

0.00

No. of Live Male Foetuses

Number

93

96

96

101

Mean

4.43

4.57

4.80

4.81

SD

2.58

1.89

2.12

2.52

No. of Live Female Foetuses

Number

83

117

83

103

Mean

3.95

5.57

4.15

4.90

SD

1.69

2.13

2.08

2.41

GROUP MEAN FOETUS WEIGHTS

 

Group

Foetus weight/Litter

(G)

Foetus Weight (G)

Male

Female

Group 1

Mean

3.3814

3.3354

3.4330

SD

0.6200

0.6853

0.5369

N

176

93

83

Group 2

Mean

3.3860

3.4788

3.3099

SD

0.5341

0.5384

0.5205

N

213

96

117

Group 3

Mean

3.3906

3.4430

3.3300

SD

0.6227

0.6619

0.5720

N

179

96

83

Group 4

Mean

3.3245

3.3427

3.3066

SD

0.7148

0.7441

0.6879

N

204

101

103

 

GROUP MEAN SEX RATIO

 

Group

Sex ratio (%) Male

Group 1

Mean

49.08

SD

17.36

N

176

Group 2

Mean

45.26

SD

16.60

N

213

Group 3

Mean

54.83

SD

17.99

N

180

Group 4

Mean

50.21

SD

17.08

N

204

 

EXTERNAL FINDINGS

 

Group

G1

G2

G3

G4

Dose (mg/kg bw)

0

100

300

1000

No. of Litter Examined

21

21

20

21

No. of Foetuses Examined

176

213

179

204

Variation Incidence – Number (%)

No. of Foetus with Variations

Total Variations

1(0.57)

6(2.82)

8(4.47)

3(1.47)

a)Foetus small in size

1(0.57)

6(2.82)

8(4.47)

3(1.47)

VISCERAL FINDINGS

 

Group

G1

G2

G3

G4

Dose (mg/kg bw)

0

100

300

1000

No. of Litter Examined

21

21

20

21

No. of Foetuses Examined

84

103

84

97

Variation Incidence – Number (%)

No. of Foetus with Variations

Total Variations

19(22.62)

18(17.48)

15(17.86)

22(22.68)

Spleen: Small in size

8(9.52)

7(6.80)

5(5.95)

7(7.22)

Spleen: Pale

5(5.95)

4(3.88)

4(4.76)

8(8.25)

Ureters: Convoluted

6(7.14)

7(6.80)

6(7.14)

7(7.22)

SKELETAL FINDINGS

Group

G1

G2

G3

G4

Dose (mg/kg bw)

0

100

300

1000

No. of Litter Examined

21

21

20

21

No. of Foetuses Examined

92

110

95

107

Variation Incidence – Number (%)

Hyoid

Not ossified

1(1.09)

0

0

0

Incompletely ossified

1(1.09)

0

0

4(3.74)

Bipartite

1(1.09)

0

0

0

Absent

1(1.09)

0

0

0

Thoracic Vertebra

Dumbbell shaped

21(22.83)

26(23.64)

18(18.95)

34(31.78)

Bipartite

3(3.26)

10(9.09)

12(12.63)

4(3.74)

Lumbar Vertebra

Not ossified

0

1(0.91)

0

0

Dumbbell shaped

0

1(0.91)

0

0

Sacral Vertebra

Not ossified

1(1.09)

0

0

0

Incompletely ossified

1(1.09)

0

0

0

Caudal Vertebra

Not ossified

6(6.52)

4(3.64)

0

5(4.67)

Sternal centers (1)

Not ossified

10(10.87)

11(10.00)

9(9.47)

12(11.21)

Incompletely ossified

10(10.87)

6(5.45)

12(12.63)

14(13.08)

Sternal centers (2)

Not ossified

5 (5.43)

3(2.73)

0

5(4.67)

Incompletely ossified

4(4.35)

0

3(3.16)

0

Misshapen/Irregular shaped

0

2(1.82)

1(1.05)

0

Manubrium

Not ossified

11 (11.96)

5(4.55)

1(1.05)

7(6.54)

Incompletely ossified

3(3.26)

0

4(4.21)

0

 

SKELETAL FINDINGS (CONTD.)

Group

G1

G2

G3

G4

Dose (mg/kg bw)

0

100

300

1000

No. of Litter Examined

21

21

20

21

No. of Foetuses Examined

92

110

95

107

Variation Incidence – Number (%)

Xyphiod

Not ossified

25 (27.17)

27(24.55)

41(43.16)

41(38.32)

Incompletely ossified

13(14.13)

17(15.45)

8(8.42)

4(3.74)

Ribs - Left

Waviness

1 (1.09)

0

0

9(8.41)

Ribs - Right

Waviness

1 (1.09)

0

0

10(9.35)

Supernumerary

0

0

0

1(0.93)

Metacarpals - Left

Not ossified

0

2(1.82)

0

1(0.93)

Incompletely ossified

1(1.09)

0

0

0

Metacarpals - Right

Not ossified

0

2(1.82)

0

1(0.93)

Incompletely ossified

1(1.09)

0

1(1.05)

0

Metatarsal - Left

Not ossified

1(1.09)

4(3.64)

0

1(0.93)

Incompletely ossified

3(3.26)

0

0

0

Metatarsal - Right

Not ossified

1(1.09)

4(3.64)

0

2(1.87)

Incompletely ossified

3(3.26)

0

0

0

Pubis - Left

Incompletely ossified

0

0

1(1.05)

0

Pubis - Right

Not ossified

0

0

1(1.05)

0

 

                                                                

Conclusions:
The No Observed Adverse Effect Level (NOAEL) of test item HOSTAPON TPHC for maternal and embryofoetal toxicity in Wistar female rats via oral route was found to be the highest dose level employed i.e., 1000 mg/kg body weight/day
Executive summary:

The test item,HOSTAPON TPHC(formulated in distilled water) was administered once daily by oral gavage to three treatment groups of twenty four pregnant female Wistar rats per group from day 5 to day 19 of gestation at dose levels of 100, 300 and 1000 mg/kg body weight/day. A control group of twenty four females were administered with vehicle (distilled water)alone.

Female animals treated at 1000mg/kg bw/day(high dose) showed burrowing followed by dullness and dyspnoea while female animals treated at 300 mg/kg bw/day (intermediate dose) showed only dullness shortly after each administration. These findings recovered to normal after 3 hours post dosing each day. These temporary effects can be regarded unspecific in nature due to minor local irritative effects caused by test item administration.

The evaluation of the reproductive organs of the dam, pre and post implantation loss, gravid uterine weight, litter size, viability of the foetus, litter weight and foetus weight, sex ratio of the foetus and the external, visceral and skeletal examination of the foetus revealed that HOSTAPON TPHC did not cause any maternal as well as embryo foetal toxicity up to the highest dose of 1000 mg/kg bw/day tested.

Effect on developmental toxicity: via oral route
Endpoint conclusion:
no study available
Quality of whole database:
The available data base is considered sufficient for hazard / risk characterization. There are no indications of developmental toxicity.

Justification for classification or non-classification

Administration of the registered substance up to the limit dose of 1000 mg/kg body weight per day did not result in statistical significant changes of reproductive and/or repeated dose toxicological parameters. Based hereupon classification and labelling of the registered substance concerning reproductive toxicity is not warranted. However, based on the general REACH information requirements, an OECD 414 prenatal developmental toxicity study in rats is included in this dossier.

Additional information

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