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Key value for chemical safety assessment

Effects on fertility

Description of key information

Based on the available data from the OECD 421 study, fertility and reproductive performance of the parental animals were adversely affected by treatment with the test substance. In addition, also developmental toxicity was evident in the offspring.

Effect on fertility: via oral route
Endpoint conclusion:
adverse effect observed
Dose descriptor:
LOAEL
30 mg/kg bw/day
Study duration:
subacute
Species:
rat
Effect on fertility: via inhalation route
Endpoint conclusion:
no study available
Effect on fertility: via dermal route
Endpoint conclusion:
no study available
Additional information

The test substance was given daily as an oily solution to groups of 10 male and 10 female Wistar rats (F0 animals) by stomach tube at doses of 30, 100 and 300 mg/kg body weight/day (mg/kg bw/d).Control animals (10 male and 10 female Wistar rats) were dosed daily with the vehicle only (corn oil). The study was conducted according to OECD 421 guideline and GLP (BASF, 2014) with some additional examinations. The duration of treatment covered a 2-week premating and mating period in both sexes, about three weeks postmating in males, and the entire gestation period as well as approximately 4 days of the lactation period in females with litters, and 4 weeks of postmating period in non-pregnant females.

 

Observations

After 2 weeks of premating treatment the F0 animals were mated to produce F1 generation pups. Mating pairs were from the same test group. Mating was discontinued as soon as sperm was detected in the vaginal smear. F0 animals were examined for their reproductive performance including determination of the number of implantation sites and the calculation of postimplantation loss for all F0 females.

 

Food consumption of the F0 parents was determined once weekly during premating. In dams food consumption was determined for gestation days 0 - 7, 7 - 14, 14 - 20 and lactation days 1 - 4.

 

Body weights of F0 parents were determined once a week, in males throughout the study and in females during premating. During gestation and lactation period, F0 females were weighed on gestation days (GD) 0, 7, 14 and 20, on the day of parturition (postnatal day [PND] 0) and on PND 4.

 

The pups were sexed and examined for macroscopically evident changes on PND 0. They were weighed on PND 1 and on PND 4. Their viability was recorded. At necropsy on PND 4, all pups were sacrificed with CO2, under isoflurane anesthesia, and examined macroscopically for external and visceral findings.

 

Clinico-chemical and hematological examinations were performed of allanimals in each sex and dose-grouptowards the end of the administration period.

 

Various sperm parameters (motility, sperm head count, morphology) were assessed in the F0 generation males at scheduled sacrifice or after appropriate staining.

 

All F0 parental animals were sacrificed by decapitation,under isoflurane anesthesia, and were assessed by gross pathology. Weights of selected organs were recorded and a histopathological examination was performed.


Results

Analysis

The various analyses:

Demonstrated the stability of the test substance in corn oil over a period of 7 days at room temperature.

- Verified correct concentrations of the test substance preparations.

 

Effects

The following test substance-related adverse effects/findings were noted:

 

300mg/kg bw/d

F0 PARENTAL ANIMALS

CLINICAL EXAMINATIONS/ REPRODUCTIVE PERFORMANCE/CLINICAL PATHOLOGY/ PATHOLOGY

- Decreased food consumption in the males during the whole premating period (up to 24% below control)

- Decreased body weights in the males on premating days 7 - 13 (up to 7% below control), during the whole mating period (about 7% below control) and during the whole postmating period (up to 13% below control), reduced body weight gain in accordance

- Indications of regenerative anemia, such as decreased red blood cell (RBC) counts, hemoglobin and hematocrit values, as well as increased relative reticulocyte counts in rats of both sexes

- Prolonged prothombin time in males

- Decreased mean corpuscular volume (MCV) and increased mean corpuscular hemoglobin concentration (MCHC) in females

- Indications ofaltered liver cell metabolism such as increased alanine aminotransferase (ALT) activities in rats of both sexes, increased creatinine values and decreased cholesterol values in males, increased triglyceride and total bilirubin values in females

- No copulation in 2/10 pairs, decreased mating index (80 vs. 100% in control)

- No pregnancies at all in females with confirmed copulation (8/10)

- Increased incidences of abnormal sperm and decreased total sperm head counts (-48 %) in the cauda epididymidis

- Decrease in absolute weight of testes (-28%), epididymides (-38%), prostate (-65%) and seminal vesicle (-84%) in male animals

- Decrease in relative weight of the epididymides (-29%), prostate (-60%) and seminal vesicle (-81%) in male animals

- Decrease in absolute (-48%) and relative (-46%) ovary weight in female animals

- Increase in absolute adrenal gland weight (+51%) in males and absolute liver weight (+14%) in female animals

- Increase in relative weight of adrenal glands (+75%/+21%), liver (+22%/+29%), and pituitary gland (+32%/+26%) of male and female animals

- Macroscopically reduced size of eight testes and epididymides and nine prostates and seminal vesicles in male animals

- Macroscopically reduced size of nine ovaries in female animals

- Moderate tubular degeneration in two males and slight reduction of tubular size in the left testis in seven males

- Slight to extreme oligospermia in four males and minimal to slight debris in seminiferous tubules in the left epididymis in three males

- Minimal to moderate increase of atretic follicles in eight females, inactivity of interstitial cells in the ovaries in six females, increased number of cyst(s) in nine females, and decreased number of corpora lutea in nine females

F1 PUPS

CLINICAL EXAMINATIONS/ GROSS FINDINGS

- No pregnancies, no pups delivered

 

100 mg/kg bw/d

F0 PARENTAL ANIMALS

CLINICAL EXAMINATIONS/ REPRODUCTIVE PERFORMANCE/CLINICAL PATHOLOGY/ PATHOLOGY

- Decreased total sperm head counts (-23 %) in the cauda epididymidis

- Decrease in absolute weight of the epididymides (-13%), prostate (-35%) and seminal vesicle (-47%) in male animals

- Decrease in relative weight of prostate (-32%) and seminal vesicle (-44%) in male animals

- Increase in relative weight of pituitary gland (+26%) of male animals

- Macroscopically reduced size of two prostates and seminal vesicles in male animals

- Minimal to slight reduction of tubular size in the left testis in six males and minimal debris in seminiferous tubules in the left epididymis in one male

- Moderate increase of atretic follicles and decreased number of corpora lutea in the ovaries in one female

- Decreased number of implantation sites (5.9** [**:p≤0.01] vs. 12.3 implants in control)

- Increased post-implantation loss (46.5% vs. 3.9% in control)

F1 PUPS

CLINICAL EXAMINATIONS/ GROSS FINDINGS

- Decreased mean number of delivered pups (4.1** [**:p≤0.01] vs. 11.8 pups per litter)

- Increased number of dead/cannibalized pups (6 vs. 1 in control), resulting in decreased viability index (about 14% below control)

- Skew of sex ratio towards female sex (33.3% males vs. 66.7% females on PND 0)

 

30 mg/kg bw/d

F0 PARENTAL ANIMALS

CLINICAL EXAMINATIONS/ REPRODUCTIVE PERFORMANCE/CLINICAL PATHOLOGY/ PATHOLOGY

- Decreased total sperm head counts (- 13 %) in the cauda epididymidis

- Decrease in absolute and relative weight of the seminal vesicle (-16%/-18%) and increase in relative pituitary gland weight (+17%)

 

 

F1 PUPS

CLINICAL EXAMINATIONS/ GROSS FINDINGS

- No test substance-related adverse findings

 

Conclusion

Under the conditions of this OECD 421 combined repeated dose toxicity study with the reproduction/developmental toxicity screening test in Wistar rats the following NOAEL (no observed adverse effect level) were determined:

 

TheNOAEL for general, systemic toxicity was 100 mg/kg bw/d for the F0 males and females based on anemia and altered liver cell metabolism at 300 mg/kg bw/d.

 

No NOAEL for fertility was determined for the F0 parental males because a dose-dependent decrease in the number of sperm in thecauda epididymidis as well as a change in relative organ weights of seminal vesicles and pituitary glands were noted at all tested dose levels of 30 mg/kg bw/d and above. Fertility and reproductive performance of F0 parental females were unaffected at 30 mg/kg bw/d. Fertility and reproductive performance were distinctly impaired at 100 mg/kg bw/d, while outright infertility was the result of the exposure to 300 mg/kg bw/d. A putative NOAEL for fertility was set at 10 mg/kg bw/d (see discussion below).

 

The NOAEL for developmental toxicity in the F1 offspring was 30 mg/kg bw/d, based on fetal and pup mortality at 100 mg/kg bw/d.

In order to establish a putative NOAEL for DNEL derivation, a safety factor of 3 (see discussion of DNEL derivation in IUCLID and chemical safety report) was chosen for the following reasons:

 

-         The sperm head counts in the cauda epididymidis were within the biological variance of the Wistar rat obtained in the laboratory conducting the study. They ranged from a mean of 298 to 688 (TS/gT [Mio/g]) in the data available from more than 30 control groups where this parameter was examined in the years from 2008 to 2013. Thus both the count of the low (422) and mid dose (372) of this study (control: 486) are within this biological range and the decreased sperm head counts in the cauda epididymidis had no effect on the pregnancy outcomes of the low (9/10 females pregnant) and mid dose (10/10 females pregnant) group.

 

-         The decreased absolute weight of the seminal vesicles (mean: 1.56 g in the control group males vs. mean: 1.307* g (*: p<= 0.05) in the low dose group males) were within the biological variance of the Wistar rat obtained in the laboratory conducting the study. They ranged from a mean of 0.905 g to 1.454 g taking into account the values of more than 40 control groups from the years 2006 to 2016. These controls include three OECD 421/422 studies where the controls ranged from 1.293 g to 1.402 g. The absolute weights of the seminal vesicles in the controls of the current study were thus rather high.

 

-         The decreased relative weight of the seminal vesicles (mean %: 0.38 in the control group males vs. mean %: 0.311** (**: p<= 0.01) in the low dose group males) was within the biological variance of the Wistar rat obtained in the laboratory conducting the study. They ranged from a mean of 0.263 % to 0.39 % taking into account the values of more than 40 control groups from the years 2006 to 2016. This includes three OECD 421/422 studies where the controls ranged from 0.334 % to 0.366 %.

 

-         The increase in relative pituitary gland weight (mean %: 0.002 in the control group males vs. mean %: 0.003* (*: p<= 0.05) in the low dose group males) was within the biological variance of the Wistar rat obtained in the laboratory conducting the study. They ranged from a mean of 0.002 % to 0.004 % taking into account the values of more than 40 control groups from the years 2006 to 2016.

 

Based on the information presented above we believe, that the safety factor of 3 is sufficiently conservative to derive the putative NOAEL for DNEL derivation. Thus the putative NOAEL for fertility is set at 10 mg/kg bw/d.

Effects on developmental toxicity

Description of key information
Based on the available data from the OECD 421 study, fertility and reproductive performance of the parental animals were adversely affected by treatment with the test substance. In addition, also developmental toxicity was evident in the offspring. 
Effect on developmental toxicity: via oral route
Endpoint conclusion:
adverse effect observed
Dose descriptor:
NOAEL
30 mg/kg bw/day
Study duration:
subacute
Species:
rat
Effect on developmental toxicity: via inhalation route
Endpoint conclusion:
no study available
Effect on developmental toxicity: via dermal route
Endpoint conclusion:
no study available
Additional information

Under the conditions of this OECD 421 combined repeated dose toxicity study with the reproduction/developmental toxicity screening test in Wistar rats the following NOAEL (no observed adverse effect level) were determined:

 

TheNOAEL for general, systemic toxicity was 100 mg/kg bw/d for the F0 males and females based on anemia and altered liver cell metabolism at 300 mg/kg bw/d.

 

No NOAEL for fertility was determined for the F0 parental males because a dose-dependent decrease in the number of sperm in thecauda epididymidis as well as a change in relative organ weights of seminal vesicles and pituitary glands were noted at all tested dose levels of 30 mg/kg bw/d and above. Fertility and reproductive performance of F0 parental females were unaffected at 30 mg/kg bw/d. Fertility and reproductive performance were distinctly impaired at 100 mg/kg bw/d, while outright infertility was the result of the exposure to 300 mg/kg bw/d.

 

The NOAEL for developmental toxicity in the F1 offspring was 30 mg/kg bw/d, based on fetal and pup mortality at 100 mg/kg bw/d.

Justification for classification or non-classification

Based on the available OECD 421 study, a classification with R60 (May impair ferility, Cat. 2) and R61 (May cause harm to the unborn child, Cat. 2) according to DSD-DPD and a classification as toxic to reproduction with Cat. 1B Fertility and Development (H360) according to CLP is warranted.