Registration Dossier

Toxicological information

Toxicity to reproduction

Currently viewing:

Administrative data

Endpoint:
screening for reproductive / developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Remarks:
OECD Test Guideline study under GLP condition
Cross-reference
Reason / purpose:
reference to same study
Reference
Endpoint:
repeated dose toxicity: oral
Remarks:
combined repeated dose and reproduction / developmental screening
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Remarks:
OECD Guidelines
Reason / purpose:
reference to same study
Qualifier:
according to
Guideline:
other: OECD TG 421: Reproduction/developmental toxicity screening test
Deviations:
no
GLP compliance:
yes
Limit test:
no
Species:
rat
Strain:
other: Crl: CD(SD)
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories Japan, Inc. Tsukuba
- Age at study initiation: 10 weeks
- Weight at study initiation: Males: 389-449 g; Females: 234-271 g
- Housing: Steel wire-mesh cage (250 mm x 350 mm x 200 mm )
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: 14 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-25
- Humidity (%): 41-69
- Air changes: 10-15 times / hr
- Photoperiod: 12 hrs dark / 12 hrs light (07:00-19:00)
Route of administration:
oral: gavage
Vehicle:
corn oil
Analytical verification of doses or concentrations:
yes
Duration of treatment / exposure:
(P) Males: 42 days including 14 days pre-mating and mating periods, and thereafter 14 days (P) Females: 42-45 days including 14 days pre-mating, mating and gestation periods, and the days until day 4 of lactation
Frequency of treatment:
Once/day, 7days/week
Remarks:
Doses / Concentrations:
0 (vehicle), 40, 200, and 1000 mg/kg bw/day
Basis:
actual ingested
No. of animals per sex per dose:
12 animals/sex/dose
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: Doses in this test were set based on the results of the following study: 28-day repeated dose oral toxicity test (doses: 0, 8, 40, 200, and 1000 mg/kg bw/day). At 1,000 mg/kg bw/day, transient salivation and tremors were observed in both sexes, decreases in body weight gain and grip strength of forearms were observed in males, and a decrease in the locomotor activity was observed in females. At this dose, increases in serum aspartate aminotransferase and alanine aminotransferase levels were observed in both sexes, whereas decreases in total protein and β-globulin fraction were observed in males, and increases in alkaline phosphatase and triglyceride levels were observed in females. Increased liver weight and decreased ovary weight were also observed at 1,000 mg/kg bw/day in females. Histopathological examinations revealed hyperostosis metaphysis of the femur in females at 200 mg/kg bw/day. Furthermore, at 1,000 mg/kg bw/day, hyperkeratosis of the mucosal epithelium in the forestomach, grade enhancement of regeneration of the tubular epithelium, dilatation of the renal tubules in the cortex, and cyst-like extension of the collecting duct in the kidney, and hyperostosis metaphysis of the femur were observed in both sexes. Additionally in males at 1,000 mg/kg bw/day, degeneration of nerve fibers in the sciatic nerve and atrophy of muscle fibers in the skeletal muscle were observed. And in females at 1,000 mg/kg bw/day, squamous cell hyperplasia of the boundary edge in the stomach was observed. These changes tended to resolve after the recovery period.
- Rationale for animal assignment (if not random): Body weight-balanced randomization
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule:
Males and females: 3 times/day

BODY WEIGHT: Yes
- Time schedule for examinations:
Males: Days 1, 4, 8, 11, 15, 22, 25, 29, 32, 36, 39, 42, and the day of necropsy
Females: Twice a week during the precopulation period (days 1, 4, 8, 11, and 15); gestation days 0, 4, 7, 11, 14, 17, and 20; lactation days 0 and 4; and the day of necropsy

FOOD CONSUMPTION: Yes
Males: Days 1, 4, 8, 11, 15, 32, 36, 39, and 42 in dosing period
Females: Days 1, 4, 8, 11, and 15; gestation days 1, 4, 7, 11, 14, 17, and 20; lactation days 2 and 4

HAEMATOLOGY: No

CLINICAL CHEMISTRY: No

URINALYSIS: No
Sacrifice and pathology:
GROSS PATHOLOGY: Yes (see tables)
HISTOPATHOLOGY: Yes (epididymis, prostate, seminal vesicle, testis, ovary, uterus, vagina, and gross abnormal sites)
Other examinations:
Organ weight: Testes and epididymides
Statistics:
The data were analyzed for homogeneity of variance by the Bartlett test. If variances were homogeneous, data was analyzed by the Dunnett test, whereas heterogeneous data was analyzed by the steel test (p<0.05, two-sided).



Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
Nine males and seven females died with tremors and decreased locomotor activity at 1,000 mg/kg bw/day by day 9 of administration.
Mortality:
mortality observed, treatment-related
Description (incidence):
Nine males and seven females died with tremors and decreased locomotor activity at 1,000 mg/kg bw/day by day 9 of administration.
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
Low value of body weight was observed in both sexes at 1,000 mg/kg bw/day.
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
Low value of food consumption was observed in both sexes at 1,000 mg/kg bw/day.
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):
not examined
Organ weight findings including organ / body weight ratios:
no effects observed
Gross pathological findings:
effects observed, treatment-related
Description (incidence and severity):
see tables.
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
see tables.
Histopathological findings: neoplastic:
not examined
Dose descriptor:
NOAEL
Effect level:
200 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: Nine males and seven females died with tremors and decreased locomotor activity at 1,000 mg/kg bw/day by day 9 of administration.
Critical effects observed:
not specified
Conclusions:
In this study, NOAEL for repeated-dose toxicity was determined to be 200 mg/kg bw/day in male and female rats.
Executive summary:

A reproduction/developmental toxicity screening test was performed according to OECD TG 421. Male and female rats (12 animals/sex/dose) were administered 4-chlorobenzaldehyde at 0, 40, 200, and 1,000 mg/kg bw/day. Males were dosed for 42 days, including a 14 day pre-mating and mating periods. Females were dosed for 42–45 days, including a 14-day pre-mating, mating, and gestation periods and the time until day 4 of lactation. Nine males and seven females died with tremors and decreased locomotor activity at 1,000 mg/kg bw/day by day 9 of administration. In this study, NOAEL for repeated-dose toxicity was determined to be 200 mg/kg bw/day in male and female rats.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2009

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
other: OECD TG 421: Reproduction/developmental toxicity screening test
Deviations:
no
GLP compliance:
yes
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): 4-chlorobenzaldehyde
- Purity: 99.8%
- Lot/batch No.: QJ5LK
- Stability under test conditions: Stable
- Storage condition of test material: a cool (2-8 °C) and dark place (in a refrigerator), with an airtight stopper
- Dosing solution storage condition: under room temperature (19-23 °C) in a brown glass bottle
- Other: The dosing solution was used within 7 days of preparation.

Test animals

Species:
rat
Strain:
other: Crl:CD(SD)
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories Japan, Inc. Tsukuba
- Age at study initiation: 10 weeks
- Weight at study initiation: Males: 389-449 g; Females: 234-271 g
- Housing: Steel wire-mesh cage (250 mm x 350 mm x 200 mm )
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: 14 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-25
- Humidity (%): 41-69
- Air changes: 10-15 times / hr
- Photoperiod: 12 hrs dark / 12 hrs light (07:00-19:00)

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
corn oil
Details on mating procedure:
- M/F ratio per cage:1:1
- Length of cohabitation:up to 14 days
- Proof of pregnancy: [vaginal plug / sperm in vaginal smear] referred to as [day 0] of pregnancy

Analytical verification of doses or concentrations:
yes
Duration of treatment / exposure:
(P) Males: 42 days including 14 days pre-mating and mating periods, and thereafter 14 days (P) Females: 42-45 days including 14 days pre-mating, mating and gestation periods, and the days until day 4 of lactation
Frequency of treatment:
Once/day, 7days/week
Doses / concentrations
Remarks:
Doses / Concentrations:
0 (vehicle), 40, 200, and 1000 mg/kg bw/day
Basis:
actual ingested
No. of animals per sex per dose:
12 animals/sex/dose
Control animals:
yes, concurrent vehicle

Examinations

Parental animals: Observations and examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule:
Males and females: 3 times/day

BODY WEIGHT: Yes
- Time schedule for examinations:
Males: Days 1, 4, 8, 11, 15, 22, 25, 29, 32, 36, 39, 42, and the day of necropsy
Females: Twice a week during the precopulation period (days 1, 4, 8, 11, and 15); gestation days 0, 4, 7, 11, 14, 17, and 20; lactation days 0 and 4; and the day of necropsy

FOOD CONSUMPTION: Yes
Males: Days 1, 4, 8, 11, 15, 32, 36, 39, and 42 in dosing period
Females: Days 1, 4, 8, 11, and 15; gestation days 1, 4, 7, 11, 14, 17, and 20; lactation days 2 and 4

OTHER: Females: Numbers of corpus luteum and implantation site on the day of necropsy
Oestrous cyclicity (parental animals):
Vaginal smears were collected from all females in the main groups and microscopically examined every day from the day after the start of administration until the day copulation was confirmed. During the pre-mating administration period, vaginal smear pictures were classified as proestrus, estrus, metestrus or diestrus and examined for the frequency of estrus and interval between estruses (estrous cycle). During the mating period, vaginal smears were examined for the presence of sperm.
Sperm parameters (parental animals):
Parameters examined in P male parental generations: testes weight, epididymides weight
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, and weight].
GROSS EXAMINATION OF DEAD PUPS: Yes, for external and internal abnormalities.
Postmortem examinations (parental animals):
SACRIFICE:
Male animals: Rats were euthanized by exsanguination under ether anesthesia on the day after the last administration.
Maternal animals: Rats were euthanized by exsanguination under ether anesthesia on day 4 of lactation.

GROSS PATHOLOGY: Yes (see tables)
HISTOPATHOLOGY: Yes (epididymis, prostate, seminal vesicle, testis, ovary, uterus, vagina, and gross abnormal sites)
ORGAN WEIGHTS, Yes: Testes and epididymis
Postmortem examinations (offspring):
GROSS NECROPSY
- Gross necropsy consisted of external and internal examinations including the cervical, thoracic, and abdominal viscera.
Statistics:
The data were analyzed for homogeneity of variance by the Bartlett test. If variances were homogeneous, data was analyzed by the Dunnett test, whereas heterogeneous data was analyzed by the Steel test (p<0.05, two-sided).
Especially,
Implantation index, Stillborn index, Liveborn index, External abnormalities, Viability index: the Steel test (p<0.05 and <0.01, two-sided)
Copulation index, Fertility index, Insemination index, Delivery index: Fisher's exact test (p<0.05 and <0.01, two-sided)
Reproductive indices:
Each parameter was determined by the following equations:
Copulation index (%) = (No. of copulated animals/No. of co-housed animals) × 100
Fertility index (%) = (No. of pregnant females/No. of copulated females) × 100
Insemination index (%) = (No. of pregnant females/No. of copulated males) × 100
Duration of gestation (days) = day 0 of lactation – day 0 of gestation
Delivery index (%) = (No. of females delivered liveborn pups/No. of pregnant females) × 100
Implantation index (%) = (No. of implantation sites/No. of corpora lutea) × 100
Stillborn index (%) = (No. of stillborn pups/Total No. of pups born) × 100
Liveborn index (%) = (No. of liveborn pups/Total No. of pups born) × 100
External abnormalities (%) = (No. of pups with external abnormalities/No. of liveborn pups) × 100
Sex ratio = No. of liveborn male pups/(No. of liveborn male pups + No. of liveborn female pups)



Offspring viability indices:
Viability index (%) = (No. of surviving pus on day 4 after birth/No. of liveborn pups on day 0 after birth) × 100

Results and discussion

Results: P0 (first parental generation)

General toxicity (P0)

Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
see 7.5.1 Repeated dose toxicity
Mortality:
mortality observed, treatment-related
Description (incidence):
see 7.5.1 Repeated dose toxicity
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
see 7.5.1 Repeated dose toxicity
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
see 7.5.1 Repeated dose toxicity
Organ weight findings including organ / body weight ratios:
no effects observed
Histopathological findings: non-neoplastic:
no effects observed
Description (incidence and severity):
on reproductive organs

Reproductive function / performance (P0)

Reproductive function: oestrous cycle:
no effects observed
Reproductive function: sperm measures:
not examined
Reproductive performance:
no effects observed
Description (incidence and severity):
on reproductive organs

Effect levels (P0)

Dose descriptor:
NOAEL
Effect level:
200 mg/kg bw/day (actual dose received)
Sex:
male/female
Basis for effect level:
other: Nine males and seven females died with tremors and decreased locomotor activity at 1,000 mg/kg bw/day by day 9 of administration.

Results: F1 generation

General toxicity (F1)

Clinical signs:
no effects observed
Mortality / viability:
no mortality observed
Body weight and weight changes:
no effects observed
Sexual maturation:
not examined
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
no effects observed
Histopathological findings:
not examined

Effect levels (F1)

Dose descriptor:
NOAEL
Generation:
F1
Effect level:
200 mg/kg bw/day (actual dose received)
Sex:
male/female
Basis for effect level:
other: Administration of 4-chlorobenzaldehyde at 1,000 mg/kg bw/day was halted because of the frequent deaths in male and female rats.

Overall reproductive toxicity

Reproductive effects observed:
not specified

Applicant's summary and conclusion

Conclusions:
NOAEL for the rat reproductive/developmental toxicity of 4-chlorobenzaldehyde was determined to be 200 mg/kg bw/day.

Executive summary:

In the reproduction/developmental toxicity screening test (0, 40, 200, and 1,000 mg/kg bw/day) (OECD TG 421), administration of 4-chlorobenzaldehyde at 1,000 mg/kg bw/day was halted because of the frequent deaths in male and female rats. No effects of this substance on reproductive and developmental parameters were observed at 200 mg/kg bw/day. NOAEL for the rat reproductive/developmental toxicity of 4-chlorobenzaldehyde was determined to be 200 mg/kg bw/day.