Registration Dossier

Administrative data

Endpoint:
two-generation reproductive toxicity
Remarks:
based on test type (migrated information)
Type of information:
experimental study
Adequacy of study:
key study
Study period:
27 May 2005 - 10 Aug 2007
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2007
Report Date:
2007

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 416 (Two-Generation Reproduction Toxicity Study)
Deviations:
no
Qualifier:
according to
Guideline:
EPA OPPTS 870.3800 (Reproduction and Fertility Effects)
Deviations:
no
Qualifier:
according to
Guideline:
other: MAFF Japan, 12-Nousan No. 8147, 2-1-17
Deviations:
no
GLP compliance:
yes
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Type:
Constituent

Test animals

Species:
rat
Strain:
other: SPF Wistar Hannover, BrlHan:WIST@Jcl[GALAS]
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Fuji Breeding Centre, CLEA Japan, Inc. (Shizuoka, Japan)
- Age at study initiation: (P) 5 weeks; (F1) app. 4 weeks
- Weight at study initiation: (P) males: 146 - 168 g, females: 110 - 125 g; (F1) males: 98 - 109 g, females: 58 - 62 g
- Fasting period before study: No
- Housing: During the pre-mating growth period, animals were housed individually in a wire-mesh stainless steel cage. Cages were arranged in an aluminum cage rack with 16 cages per rack. A steel cage tray was placed under each cage. Racks and cages were exchanged for sterile cages once every 4 weeks and cage trays were exchanged twice weekly. A male and a female of the same group were housed together in an aluminium cage with wire-mesh floor and front and a cage tray (W260 x D400 x H240 mm) for mating. Cages were arranged in stainless steel cage racks with 20 cages per rack. Females were housed individually in aluminium breeding boxes ((W260 x D400 x H200 mm) with nesting material (Sunflake: Oriental Yeast Co., Ltd., Tokyo, Japan) for delivery and nursing during the gestation and lactation period. Offspring were housed per litter in the breeding boxes after weaning and the boxes were set on stainless steel racks (20 boxes/rack). Males that accomplished mating and females that accomplished lactation were housed until necropsy in the same way as described for the pre-mating period.
- Diet: Certified pulverized feed (MF Mash, Oriental Yeast Co.,Ltd., Tokyo, Japan) ad libitum
- Water: Filtered and sterilised well water, ad libitum
- Acclimation period: 7 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ±2
- Humidity (%): 55±15
- Air changes (per hr): > 10 per hour
- Photoperiod (hrs dark / hrs light): 12/12 hrs

IN-LIFE DATES: From: app. 01 Jun 2005 To: 17 Feb 2006

Administration / exposure

Route of administration:
oral: feed
Vehicle:
unchanged (no vehicle)
Details on exposure:
PREPARATION OF DOSING SOLUTIONS:

DIET PREPARATION
- Rate of preparation of diet (frequency): Every 1 - 4 weeks, in total 10 times during the test period
- Mixing appropriate amounts with: Test substance was mixed in a mortar with a small amount of certified pulverised feed; this mixture was then mixed with the feed in a blending machine.
- Storage temperature of food: 1 - 10 °C
Details on mating procedure:
- M/F ratio per cage: 1 male and 1 female per cage during the mating period.
- Length of cohabitation: From 1 up to 14 days, until indication of copulation was found in the female.
- Proof of pregnancy: Vaginal plug or sperm in vaginal smear; referred to as day 0 of pregnancy.
- After 14 days of unsuccessful pairing replacement of first male by another male with proven fertility. The mating period continued up to 1 additional week.
- Further matings after two unsuccessful attempts: No
- After successful mating each pregnant female was caged (how): Females were housed individually in aluminium breeding boxes ((W260 x D400 x H200 mm) with nesting material (Sunflake: Oriental Yeast Co., Ltd., Tokyo, Japan) for delivery and nursing during the gestation and lactation period.
- Any other deviations from standard protocol: No
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Chemical analyses for homogeneity and concentration were performed on the first and sixth preparation of the feed on samples taken from the top, middle and bottom layer of the mixer for each dose level.
Duration of treatment / exposure:
P: 10-week pre-mating period and during the mating-, gestation- and lactation periods for breeding the F1 litters (8 weeks for males, 8 - 10 weeks for females).
F1: Maintained on their respective diets from the day of weaning (day 21 post partum), during their growth into adulthood (10 weeks pre-mating period) and also during the mating-, gestation- and lactation periods for breeding the F2 litters (8 weeks for males, 8 - 10 weeks for females).
Frequency of treatment:
Daily
Details on study schedule:
- F1 parental animals not mated until app. 6 weeks after selection from the F1 litters
- Selection of parents from F1 generation when pups were 21 - 25 days of age
- Age at mating of the mated animals in the study: App. 10 weeks
Doses / concentrations
Remarks:
Doses / Concentrations:
0, 2.97, 59.7, 590 mg/kg bw/d (males) and 0, 4.87, 97.6, 976 mg/kg bw/d (females)
Basis:
actual ingested
No. of animals per sex per dose:
P: 24
F1: 24
Control animals:
yes, plain diet
Details on study design:
- Dose selection rationale: A dose-range finding study was performed with 8 rats/sex/group, administered 0, 600, 2000, 6000 and 20000 ppm for a period of 3 weeks prior to mating and until weaning of the F1 offspring. Systemic toxicity (statistically significant increase in absolute and relative liver weight) was reported in the 2000 ppm and above for males, and the 6000 and 20000 ppm groups for females in the P generation. Pup weights were retarded in the 6000 and 20000 ppm group for males, and in the 20000 ppm group for females. The absolute and relative thymus weight of the pups in the 20000 ppm dose group were statistically significantly decreased compared to the control group.
Positive control:
No

Examinations

Parental animals: Observations and examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Twice daily
- Cage side observations included: Clinical signs, mortality

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: A record of mating of the females was made by examination of the daily vaginal smears for spermatozoa and/or appearance of a vaginal plug throughout the mating periods. Towards the end of the gestation period, females were examined twice daily for signs of parturition. Duration of the gestation was calculated. As soon as possible after parturition, the litters were examined for litter size, live birth, stillbirth and gross anomalies. The sex ratio of pups was recorded on day 0 of lactation. Pups were weight individually on days 0, 4, 7, 14 and 21 of lactation. The number of live pups in each litter was counted on days 0, 4, 7, 14 and 21 of the lactation period. The dams and pups were observed daily for survival and behavioral abnormalities in nesting and nursing.

BODY WEIGHT: Yes
- Time schedule for examinations: The body weights of males and females were recorded at weekly intervals (with exception of the pairing periods). After mating, females were weighed on days 0, 7, 14 and 20 of gestation. Dams which littered were weighed on days 1, 4, 7, 14 and 21 post partum.


FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study):
The food consumption was recorded weekly, together with the recording of the body weights with exception of the pairing periods. The relative food consumption ratios and intake of the test article expressed per mg/kg/day were calculated.
- 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: No
Sperm parameters (parental animals):
Parameters examined in P/F1 male parental generations: Testis weight, epididymis weight, sperm head count in testes, sperm count in epididymides, sperm motility, sperm morphology.
Litter observations:
STANDARDISATION OF LITTERS
- Performed on day 4 postpartum: Yes
- If yes, maximum of 8 pups/litter (4/sex/litter as nearly as possible); excess pups were killed and discarded

PARAMETERS EXAMINED
The following parameters were examined in F1 / F2 offspring: Number and sex of pups, stillbirths, live births, postnatal mortality, presence of gross anomalies, weight gain, physical or behavioural abnormalities.

GROSS EXAMINATION OF DEAD PUPS:
Yes, for external and internal abnormalities; possible cause of death was determined for pups born or found dead.
Postmortem examinations (parental animals):
SACRIFICE
- Male animals: All surviving animals, week 18 of the exposure period
- Maternal animals: All surviving animals, after the last litter of each generation was weaned (week 18-20 of the exposure period)

GROSS NECROPSY
From all P and F1 animals selected for pairing, samples of the following tissues and organs were collected at necropsy and fixed in neutral-buffered 10 % formalin solution: gross lesions, adrenals, brain (incl. entire brainstem), epididymides, heart, kidneys, liver, ovaries, prostate, pituitary, seminal vesicles with fluids, thyroid, spleen, uterus (wih cervix and oviducs), vagina. Testes were fixed in a FSA (formailn-sucrose-acetic acid) solution.
External and internal examinations were performed and gross lesions recorded.

HISTOPATHOLOGY / ORGAN WEIGHTS
Organ weights: The following organ weights were recorded from all P and F1 parental animals after necropsy: adrenals, brain (incl. entire brainstem), epididymides, heart, kidneys, liver, ovaries, prostate, pituitary, seminal vesicles with fluids, thyroid, spleen, testes and uterus (with cervix and oviducs).
Organ/body weight ratios were determined.

Histopathology: Performed on 10 pairs of randomly selected high dose and control P and F1 animals, and from animals killed in extremis or which died during the study for the following organs: gross lesions, ovaries,oviducts, uterus (cornua and cervix), vagina, prostate, testes, epididymides, seminal vesicles with coagulating glands, pituitary gland and adrenals.
Additionally, from the P and F1 animals of the control and high dose group the liver was examined histologically; from F1 males in the control and high dose group: kidney, adrenals, spleen; the kidney of females in the P generation and the spleen of females in the F1 generation. The reproductive organs of infertile males and females were examined histologically from the control and the high dose groups (P and F1 animals). Organs with macroscopic abnormalities were examined histologically from the control and the high dose groups (P and F1 animals) and from the low- and mid-dose groups (P animals).
Postmortem examinations (offspring):

SACRIFICE
- Excess F1 and F2 pups after standardization of litter sizes were sacrificed on day 4 post partum and necropsied. The F1 offspring not selected as parental animals and all F2 offspring were sacrificed after weaning and necropsied. Gross pathlocial finding were recorded.
- One male and one female each of these animals were subjected to postmortem examinations (macroscopic and/or microscopic examination).

GROSS NECROPSY
From all F1 animals selected for pairing and from one male and one female pup (selected for organ weight recording) of each F1 and F2 litter, samples of the following tissues and organs were collected at necropsy and fixed in neutral buffered 10 % formaldehyde solution: gross lesions, brain, spleen, thymus, liver, ovaries, prostate, seminal vesicles, testes, epididymides, uterus and vagina.

HISTOPATHOLOGY / ORGAN WEIGTHS
Organ weights: The following organ weights were recorded from one male and one female pup of each F1 and F2 litter (in the average weight of that litter on day 14 post partum), on day 21 post partum or shortly thereafter: Brain, spleen, thymus and uterus.

From one male and one female pup (selected for organ weight recording) of high dose and control F1 and F2 litters the thymus was examined histologically. The reproductive organs of infertile males and females were examined histologically from the control and the high dose groups (F1 animals). Organs with macroscopic abnormalities were examined histologically from the control and the high dose groups (F1 adult animals and F2 pups).
Statistics:
The following statistical methods were used to analyze body weights, food consumption, organ weights, reproduction data and breeding data: Equality of variances were first evaluated by Bartlett's test (p=0.05). When group variances were homogenous, a parametric analysis of variance in one-way classificaitons (p=0.05) was used to determine statistical differences between groups. When the analysis of variance was significant, Dunnett's multiple comparison test (p=0.05 or 0.01) was perfomred to detect any statistically significant differences between treated and control groups. When Bartlett's test indicated variances were not homogenous, the Kruskal-Wallis test (p=0.05) was used to detect statistically significant differences between treated and control groups, and when significant the Dunnett-type mean rank test (p=0.05 or 0.01) was used to detect statistically significant diffetrences between treated and control group. Ovarian follicle counts were first evaluated by the F test (p=0.05) and when group variances were homogenous, Student's t-test (p=0.05 or 0.01) was performed to detect statistically significant diffetrences between treated and control groups. When variances were not homogenous, Aspin-Welch's t-test (p=0.05 or 0.01) was used. For the data on sexual devalopment, estrou cycle length, number of days until mating, duration of gestation, percentage motiliy and morphologically normal epididymes sperm form parental animals, and viability indices from offspring; the Kruskal-Wallis test (p=0.05) was used. When significant the Dunnett-type mean rank test (p=0.05 or 0.01) was used. Fisher's exact probability test (p=0.05, 0.01 or 0.001) was used for indices of clinical, gross pathological, and histopathological findings, indices of females with normal estrous cycles, and mating, fertility and gestation indices from parental animals, and sex ratios from pups. Mann-Whitney's U-test (p=0.05 or 0.01) was used (offspring, clinical, pathological indices).

Results and discussion

Results: P0 (first parental animals)

General toxicity (P0)

Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
incidental occurrences of fur loss, wounds and soiled fur in several or all dose groups; 1 female in the high dose group and 1 male in the control group were euthanised due to emaciation and abnormal respiratory sounds, respectively
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
The effects on body weight in the mid- and high dose group of the F1 males are considered to be treatment-related.
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
The effects on body weight in the mid- and high dose group of the F1 males are considered to be treatment-related.
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
P+F1 males/females: increase in liver weight (high and mid dose groups); high dose groups: F1 males: increased weights of the kidneys, spleen, adrenals; P females: increase in kidney weights; F1 females: increased weights of the adrenals and spleen
Gross pathological findings:
effects observed, treatment-related
Description (incidence and severity):
incidental finding of hair loss, enlarged thyroid, pelvic dilatation of the kidney was observed in all dose groups
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
Some incidents of microgranuloma and extramedullary hematopoiesis in the liver of one or both sexes, and extramedullary hematopoiesis in the spleen of infertile males and females.
Other effects:
no effects observed

Reproductive function / performance (P0)

Reproductive function: estrous cycle:
no effects observed
Reproductive function: sperm measures:
no effects observed
Reproductive performance:
no effects observed

Details on results (P0)

CLINICAL SIGNS AND MORTALITY (PARENTAL ANIMALS)

There were incidental occurrences of fur loss, wounds and soiled fur in several or all dose groups; therefore these were considered not to be treatment-related. One female in the 1000 ppm group was euthanised in the 5th week after exhibiting emaciation, decreased spontaneous motor activity and red discharge. One male in the control group was euthanised in the 15th week due to abnormal respiratory sounds and soiled fur.

BODY WEIGHT AND FOOD CONSUMPTION (PARENTAL ANIMALS)

Males:
Mean body weights in the P generation treated groups were comparable to those of the control group. In the F1 generation, the body weight was statistically significantly reduced from week 7 onward in the mid-dose group and throughout the study period in the high dose group. The body weight gain was statistically significantly reduced compared to the control group of the P generation week 0-2 in the mid-dose group and week 0 - 1 in the high dose group. This is not considered to be a toxicologically relevant effect, due to the brevity of the effect. In the F1 generation, a statistically significant decline in body weight gain was observed in the mid-dose group from week 6 onwards, with the exception of week 12, and in the high dose group in weeks 1-11 and week 16. The food consumption in the treated groups was comparable to the control group in the P generation. For the F1 generation, food consumption was reduced in the mid-dose group in weeks 4, 5, 6, 13 and 16, and in the high dose group in weeks 3, 4, 6, 13, and 16.
The effects on body weight in the mid- and high dose group of the F1 males are considered to be related to the treatment with the test substance.

Females:
Mean body weights and body weight gains in the P generation treated groups were comparable to those of the control group. In the F1 generation, the body weight was significantly reduced during week 2 - 4 in the high dose group. The mean body weight gain was significantly reduced in weeks 0 - 3 in the high dose group only. The food consumption was normal in P and F1 generations during the study period.

TEST SUBSTANCE INTAKE (PARENTAL ANIMALS)

Males:
Mean values for test article intake, based on food consumption and body weight determinations:
P generation, pre-mating growth period:
3.35 mg/kg body weight/day at 50 ppm
66.8 mg/kg body weight/day at 1000 ppm
662 mg/kg body weight/day at 10000 ppm

F1 generation, pre-mating growth period:
4.05 mg/kg body weight/day at 50 ppm
80.6 mg/kg body weight/day at 1000 ppm
823 mg/kg body weight/day at 10000 ppm

P generation, breeding period:
2.35 mg/kg body weight/day at 50 ppm
48.1 mg/kg body weight/day at 1000 ppm
470 mg/kg body weight/day at 10000 ppm

F1 generation, breeding period:
2.38 mg/kg body weight/day at 50 ppm
47.5 mg/kg body weight/day at 1000 ppm
482 mg/kg body weight/day at 10000 ppm

Means of P generation during the entire study (wk 1-17):
2.97 mg/kg body weight/day at 50 ppm
59.7 mg/kg body weight/day at 1000 ppm
590 mg/kg body weight/day at 10000 ppm

Means of F1 generation during the entire study (wk 1-17):
3.42 mg/kg body weight/day at 50 ppm
68.2 mg/kg body weight/day at 1000 ppm
695 mg/kg body weight/day at 10000 ppm

Females:
Mean values for test article intake, based on food consumption and body weight determinations:
P generation:
Pre-mating growth period:
4.16 mg/kg body weight/day at 50 ppm
83.9 mg/kg body weight/day at 1000 ppm
831 mg/kg body weight/day at 10000 ppm

Gestation and lactation periods:
6.05 mg/kg body weight/day at 50 ppm
120.4 mg/kg body weight/day at 1000 ppm
1217 mg/kg body weight/day at 10000 ppm

F1 generation:
Pre-mating growth period:
4.74 mg/kg body weight/day at 50 ppm
95.0 mg/kg body weight/day at 1000 ppm
941 mg/kg body weight/day at 10000 ppm

Gestation and lactation periods:
5.78 mg/kg body weight/day at 50 ppm
115 mg/kg body weight/day at 1000 ppm
1170 mg/kg body weight/day at 10000 ppm

Means of P generation during the entire study (wk 1-lactation day 21):
4.87 mg/kg body weight/day at 50 ppm
97.6 mg/kg body weight/day at 1000 ppm
976 mg/kg body weight/day at 10000 ppm

Means of F1 generation during the entire study (wk 1-lactation day 21):
5.13 mg/kg body weight/day at 50 ppm
103 mg/kg body weight/day at 1000 ppm
1027 mg/kg body weight/day at 10000 ppm

REPRODUCTIVE FUNCTION: ESTROUS CYCLE (PARENTAL ANIMALS)

No treatment-related effects on the estrous cycle of the females were observed.

REPRODUCTIVE FUNCTION: SPERM MEASURES (PARENTAL ANIMALS)

No treatment-related effects on sperm measures (testicular sperm head count, epididymal sperm number, percent sperm motility and percent normal morphology) were observed.

REPRODUCTIVE PERFORMANCE (PARENTAL ANIMALS)

The reproduction performance of the P and F1 generation animals in the dose groups (50, 1000 and 10000 ppm) - as assessed by: mating index and days until mating; fertility; gestation index; duration of gestation; mean number of implantation sites per dam; mean number of live or dead pups per dam at parturition; postnatal loss - was not considered to be affected by treatment with the test substance.

ORGAN WEIGHTS (PARENTAL ANIMALS)

Males:
The absolute and relative liver weight in the P generation was significantly increased in the mid- and high dose group, while the absolute liver weight and absolute plus relative liver weight was significantly increased in the F1 generation mid- and high dose group, respectively. In the F1 generation high dose group, the relative weights of kidneys, spleen and adrenals were also significantly increased.

Females:
The absolute and relative liver weight in the P generation was significantly increased in the mid- and high dose group, while the absolute plus relative liver weight was significantly increased in the F1 generation high dose group. Females in the high dose group also showed significant increases in the absolute and relative kidney weights of the P generation, as well as increased relative weights of the adrenals and spleen in the F1 generation.

There were no histopathological findings indicating a treatment-related effect in the liver. Therefore, the liver weight increases in both sexes is considered to be an adaptive response to the test substance. Although there were no histopathological findings in the spleen, kidneys or adrenals, the increase in spleen weight may indicate an effect on the haematopoietic system. The increase in kidney weight could be a result of increased water intake (not measured) and is, toghether with the adrenal weight increase, seen as not treatment-related.

GROSS PATHOLOGY (PARENTAL ANIMALS)

Incidental finding of hair loss, enlarged thyroid, pelvic dilatation of the kidney was observed in all dose groups. No treatment-related changes were reported.

HISTOPATHOLOGY (PARENTAL ANIMALS)

Ten pairs of animals in the control and high dose groups on the P and F1 generation, which produced weanlings, were selected for histopathological examination of the reproductive organs, pituitary gland and adrenal glands. Organs of all the animals in the control and high dose groups with statistically significant increases in their absolute and/ or relative weights apparently due to the treatment (the liver of males in the P and F1 generations; the adrenal glands, kidney and spleen of males in the F1 generation; the liver of females in the P and F1 generations; the kidneys of females in the P generation; and the spleen of females in the F1 generation) were also examined histopathologically. In addition, the reproductive organs, pituitary gland and adrenal glands of infertile animals of both sexes were subjected to histopathological examination.

The histopathological examination of the reproductive organs of fertile males revealed atrophy of the seminiferous tubules in 1 male each in the control and high dose groups of the P generation; vacuolation of epididymal tubular cells and decreased secreted materials in the coagulating glands in 1 male each in the high dose group of the P generation; and infiltration of mononuclear cells in the prostate in 1 male each in the control and high dose groups of the P generation and in 3 and 2 males in the control and high dose groups of the F1 generation, respectively. No statistically significant difference was seen between the control and dose groups for any changes.
In the reproductive organs of the infertile males, atrophy of the seminiferous tubules with epididymal oligospermia and castration cells in the pituitary in 1 control male; infiltration of mononuclear cells in the prostate in 1 male in the low dose group; focal hypertrophy of cortical cells in 1 male in the mid-dose group; and atrophy of glandular epithelial cells in prostate of 1 male in the high dose group was observed - all in the F1 generation.

No abnormalities were found in the reproductive organs, pituitary gland and adrenal glands of females, with the exception of 1 control female in the F1 generation with a cyst in the anterior lobe of the pituitary gland. In the infertile females, vaculation of vaginal mucosal epithelial cells in 1 female in the low dose group of th P generation and vaginal atresia with luminal dilatation of the uterus in 1 female in the high dose group of the F1 generation was observed. None of these findings are statistically significant.

The examination of the major organs did not show any statistically significant changes or treatment-related effects. Some incidents of microgranuloma and extramedullary hematopoiesis in the liver of one or both sexes, and extramedullary hematopoiesis in the spleen of infertile males and females. For both sexes, pelvic dilatation of the kidneys was observed in control and high dose groups, along with incidental cases of tubular basophilic changes, urinary cast, pyelitis with/without fibrosis, and hyperplasia of proximal tubular cells. None of the differences were statistically significant.

Effect levels (P0)

open allclose all
Dose descriptor:
NOAEL
Remarks:
reproductive toxicity
Effect level:
> 10 000 ppm
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: corresponding to > 590 mg/kg bw/day for males and > 976 mg/kg bw/day for females; highest dose tested
Remarks on result:
other: Generation: P and F1 adults (migrated information)
Dose descriptor:
NOAEL
Remarks:
systemic toxicity
Effect level:
> 10 000 ppm
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: corresponding to > 590 mg/kg bw/day for males and > 976 mg/kg bw/day for females; highest dose tested
Dose descriptor:
NOAEL
Remarks:
systemic toxicity
Effect level:
50 ppm
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: see 'Remark'
Remarks on result:
other: Generation: F1 and F2 pups (migrated information)

Results: F1 generation

General toxicity (F1)

Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
wound in the umbilical cord region, small body size, subcutaneous haemorrhage and fur loss; the findings were considered to be incidental
Mortality / viability:
no mortality observed
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
in the high and mid dose groups, the body weight of F1 and F2 pups of both sexes was significantly decreased
Sexual maturation:
no effects observed
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
significant decrease in thymus weight in male mid-dose F2 pups and high dose F1 and F2 pups of both sexes; as no histopathological findings were noted, the effect was considered to be not treatment-related
Gross pathological findings:
effects observed, treatment-related
Description (incidence and severity):
the incidence of dilatation of the renal pelvis was increased; this finding is not considered to be treatment-related as it is known for the used rat strain
Histopathological findings:
effects observed, treatment-related
Description (incidence and severity):
An increase in lymphophagocytosis of the thymus was observed in the control groups and treated groups in similar incidences. The findings are therefore probably not caused by the test substance.

Details on results (F1)

VIABILITY (OFFSPRING)

There was no statistically significant difference in viability (measured as viability from birth to lactation day 21) between the control and treated groups in F1 and F2 pups. The number of pups delivered was also similar between control and treated groups.

CLINICAL SIGNS (OFFSPRING)

The number of still births on lactation day 0 was not significantly different between the control and treated groups of F1 and F2 pups. Incidental clinical findings from lactation day 0-21 included wound in the umbilical cord region, small body size, subcutaneous haemorrhage and fur loss; but none of the findings were treatment-related.

BODY WEIGHT (OFFSPRING)

The mean body weight of F1 and F2 pups of both sexes in the low dose group were comparable to the control group at all time points. In the mid-dose group, the body weight of F1 pups of both sexes and of male F2 pups at lactation day 21 were significantly lower than for pups in the control group. In the high dose group, the body weight of F1 and F2 pups of both sexes was significantly decreased compared to the control group at birth and lactation day 21. In addition, significantly lower body weight was reported lactation day 4 and 14 for F1 high dose male and female pups; and lactation day 14 for F2 high dose male pups. The effects seen in the high dose group are considered to be treatment-related, but as maternal toxicity was seen at the same dose level these are not developmental effects.

SEXUAL MATURATION (OFFSPRING)

There were no treatment-related effects on the sexual maturation of the F1 and F2 pups. There was no statistically significant difference in the sex ratio of F1 and F2 pups.

ORGAN WEIGHTS (OFFSPRING)

The mean organ weights of F1 and F2 pups of both sexes in the low dose group were comparable to those of the control group. The absolute weight of the thymus was statistically significantly decreased in male mid-dose F2 pups and high dose F1 and F2 pups of both sexes. In the male pups the decrease was dose-related. The relative thymus weight was also statistically reduced in male and female F2 pups in the high dose group. The changes in the high dose group may be caused by exposure to the test substance. However, no histopathological changes were observed. Furthermore, the organ weight changes are related to the maternal toxicity observed at the same dose level.

A decrease in the absolute spleen weight in the mid-dose group of F2 pups is not considered to be treatment-related, as no effect was seen on the absolute or relative spleen weight in the high dose group of F1 and F2 pups. The observed increase in relative brain weight in male and female F1 and F2 weanlings is also considered to be incidental, as there was no difference in absolute brain weight and the body weights of all the high dose groups were significantly lower than that of the control groups. There were no effects on the uterus weight of the female F1 and F2 pups.

GROSS PATHOLOGY (OFFSPRING)

No treatment-related findings of the macroscopic examination were observed in the F1 or F2 pups in the treated groups. The number of weanlings with dilatation of the renal pelvis was 6, 2, 10 and 12 in the F1 control, low- mid- and high dose groups; and 8, 8, 10 and 24 in the F2 control, low- mid- and high dose groups, respectively. The incidence of dilatation of the renal pelvis is known to occur relatively frequently in this rat strain and the incidence was not statistically significant for any treated group compared to the control group. This is therefore not considered to be a treatment-related finding.

HISTOPATHOLOGY (OFFSPRING)

The thymus of all F1 and F2 weanling were subject to a histopathological examination due to the significant decrease in weight in several treated groups. An increase in lymphophagocytosis of the thymus was observed in the control groups and treated groups in similar incidences. The findings are therefore probably not caused by the test substance.

Effect levels (F1)

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Dose descriptor:
NOAEL
Remarks:
systemic toxicity
Generation:
F1
Effect level:
50 ppm
Based on:
test mat.
Sex:
male
Basis for effect level:
other: corresponding to 3.42 mg/kg bw/day; based on reduced body weights
Dose descriptor:
NOAEL
Remarks:
systemic toxicity
Generation:
F1
Effect level:
1 000 ppm
Based on:
test mat.
Sex:
female
Basis for effect level:
other: corresponding to 103 mg/kg bw/day; based on reduced body weights

Overall reproductive toxicity

Reproductive effects observed:
not specified

Applicant's summary and conclusion