Registration Dossier

Toxicological information

Toxicity to reproduction

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Administrative data

Endpoint:
screening for reproductive / developmental toxicity
Remarks:
based on test type (migrated information)
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Study period:
2012-10-16 to 2012-11-29 (in-life phase)
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Read-across from a guideline study with GLP, RL1

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2013
Report Date:
2013

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 421 (Reproduction / Developmental Toxicity Screening Test)
GLP compliance:
yes (incl. certificate)
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Type:
Constituent
Test material form:
solid: particulate/powder
Remarks:
migrated information: powder
Details on test material:
- Name of test material (as cited in study report): Tellurium dioxide
- Substance type: inorganic

Test animals

Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories, Research Models and Services, Germany GmbH, Sandhofer Weg 7, D-97633, from SPF colony
- Age at study initiation: Young adult rats, approximately 10 weeks old at starting and 12 weeks at mating.
- Weight at study initiation: 331-371 g males and 195-247 g females
- Fasting period before study: no data
- Housing: Type II and III polypropylene/polycarbonate, Rodents were group-housed as practical, up to 4 animals of the same group per cage
- Diet (e.g. ad libitum): ssniff® SM R/M ad libitum
- Water (e.g. ad libitum): tap water from municipal supply ad libitum
- Acclimation period: 6 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21.0 -24.8°C
- Humidity (%): 32 - 61 %
- Air changes (per hr): 15-20 air exchanges/hour
- Photoperiod (hrs dark / hrs light): light 12 hours daily, from 6.00 a.m. to 6.00 p.m.

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
other: 1% aqueous Methylcellulose
Details on exposure:
PREPARATION OF DOSING SOLUTIONS:

VEHICLE
- Justification for use and choice of vehicle (if other than water): 1% aqueous Methylcellulose (abbreviation: 1% MC) was used as vehicle.
- Concentration in vehicle:
- Amount of vehicle (if gavage): 5 mL/kg bw
- Lot/batch no. (if required): O16147824 (Dow Chemicals)
Details on mating procedure:
Mating began 2 weeks after the initiation of treatment with one female and one male of the same dose group (1:1 mating) in a single cage. Females remained with the same male during the 14-days mating period or until the copulation occurred.

A vaginal smear was prepared daily for each female during the mating period and stained with 1% aqueous methylene blue solution. The smears were examined with a light microscope, the presence of vaginal plug or sperm in the vaginal smear was considered as evidence of copulation (Day 0 of pregnancy as defined by the relevant guidelines). Sperm positive females were caged individually. Mating pairs were clearly identified in the data, mating of siblings was avoided.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Analysis of test item formulations for concentration and homogeneity was performed on 3 occasions, during the first and last weeks and approximately midway during the treatment, using a validated ICP method based on the tellurium content.
Duration of treatment / exposure:
Males were dosed for 28 days (14 days pre-mating and 14 days mating/post- mating), then they were euthanized and subjected to necropsy examination.
Females were dosed for 14 days pre-mating, for up to 14 days mating period, through gestation and up to and including the day before necropsy, 4 days post-partum dosing. The day of birth (viz. when parturition was complete) was defined as Day 0 post-partum.
Frequency of treatment:
7 days/week
Details on study schedule:
Females 4503 and 4512 showed on evidence of copulation (not-mated) and not-delivered females 4501, 4504 and 4510 were sacrificed 26 days after the last day of mating, as practical.

All F1 offspring were terminated on Day 4 post-partum.
Doses / concentrationsopen allclose all
Remarks:
Doses / Concentrations:
0 mg/kg bw/d
Basis:
actual ingested
Remarks:
Doses / Concentrations:
25 mg/kg bw/d
Basis:
actual ingested
Remarks:
Doses / Concentrations:
120 mg/kg bw/d
Basis:
actual ingested
Remarks:
Doses / Concentrations:
600 mg/kg bw/d
Basis:
actual ingested
No. of animals per sex per dose:
12 male and 12 female animals per dose group
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: The dose levels were selected at 25, 120 and 600 mg/kg bw/day at a constant dose volume of 5 mL/kg, based on available data and information from previous experimental work, including the results of a preliminary dose range finding study
- Rationale for animal assignment:
All parental (P) animals were sorted according to body weight by computer and divided to weight ranges. There were an equal number of animals from each weight group in each of the experimental groups assigned by randomisation to ensure that animals of all test groups were as nearly as practicable of a uniform weight.
Positive control:
No positive control

Examinations

Parental animals: Observations and examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: twice daily (at the beginning and end of each day)
- Cage side observations: Parental animals were inspected for signs of morbidity and mortality

DETAILED CLINICAL OBSERVATIONS: Yes
- General clinical observations were made once a day, after treatment at approximately the same time as practical. Pertinent behavioural changes, signs of difficult or prolonged parturition and all signs of toxicity including mortality were monitored.
- More detailed examinations were made once before the first exposure (to allow for within-subject comparisons), and at least once a week thereafter.
Signs evaluated included monitoring of any changes in skin, fur, eyes, mucous membranes, occurrence of secretions and excretions, and autonomic activity (e.g. lacrimation, piloerection, pupil size, unusual respiratory pattern), changes in gait, posture and response to handling as well as the presence of clonic or tonic movements, stereotypies (e.g. excessive grooming, repetitive circling), difficult or prolonged parturition or bizarre behaviour (e.g. self-mutilation, walking backwards). Special attention was directed towards the observation of tremors, convulsions, salivation, diarrhoea, lethargy, sleep and coma.

BODY WEIGHT: Yes
- Time schedule for examinations: All adult animals were weighed on Day 0, afterwards at least weekly and at termination.
Parent females were weighed on gestation Days GD0, 7, 14 and 20 and on postpartal Days PPD0 (within 24 hours after parturition) and 4
(before termination). The females were additionally weighed on GD4, 10 and 17 in order to give accurate treatment volumes.

FOOD CONSUMPTION AND COMPOUND INTAKE: Yes
Animal food consumption was determined by re-weighing the non-consumed diet with a precision of 1 g on Day 7 then at least weekly

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): No

OTHER: On gestation day GD13 the sperm positive females were examined for the presence of vaginal bleeding or “placental sign” (intrauterine extravasation of blood as an early sign of pregnancy in rat).
Estrous cyclicity (parental animals):
Detailed histological examination of the ovaries covered the follicular, luteal, and interstitial compartments of the ovary, as well as the epithelial capsule and ovarian stroma.
Sperm parameters (parental animals):
Special attention was paid to evaluation of the stages of spermatogenesis in the male gonads and histopathology of interstitial testicular cell structure.
Litter observations:
Each litter was examined as soon as possible after delivery to establish the number and sex of pups, stillbirths, live births, runts (pups that are
significantly smaller than normal pups) and the presence of gross abnormalities. Live pups were counted, sexed, weighed individually within 24 hoursof parturition (on the first day after parturition was complete, i.e. Day 0 or 1 post-partum) and on Day 4 post partum.

Observations are reported individually for each adult and offspring. All the litters were checked and recorded daily for the number of viable and dead pups. The dead pups found were subjected to necropsy with macroscopic examination. All observed abnormalities were recorded and are reported.
Some of the pups that were found dead were cannibalised, thus, they were counted and sex determined (when it was possible) but were not further
examined macroscopically.
Postmortem examinations (parental animals):
SACRIFICE
- Male animals: All surviving animals after 28 days, (14 days pre-mating and 14 days mating/post- mating)
- Maternal animals: All surviving animals were sacrificed after 4 days post partum.
Females showed on evidence of copulation (not-mated) and not-delivered females were sacrificed 26 days after the last day of mating, as practical.

GROSS NECROPSY
Gross necropsy was performed on each animal irrespective of the date of death.
After exsanguination the external appearance was examined, cranium, thoracic and abdominal cavities were opened and the appearance of the tissues and organs was observed macroscopically. Any abnormality was recorded with details of the location, colour, shape and size, as appropriate.

HISTOPATHOLOGY / ORGAN WEIGHTS
Detailed histological examination was performed on the selected list of retained organs (brain, uterus, ovaries, vagina, testes, epididymides, prostate, seminal vesicles with coagulating glands, and pituitary) in the control and High dose groups, found dead animals and any macroscopic findings (abnormalities) observed.
Special attention was paid to the organs of the reproductive system. The number of implantation sites and of corpora lutea was recorded.

At the time of termination, body weight and weight of the following organs of all parental animals were determined:
- With a precision of 0.01 g: uterus (with and without cervix), vagina, testes, epididymides, prostate, seminal vesicles with coagulating glands, brain
- With a precision of 0.001 g: ovaries, pituitary

Paired organs were weighed individually; absolute organ weights were measured and are reported. Relative organ weights (to body and brain weight) were
calculated and are reported.
The weighed organs and all organs showing macroscopic lesions of all adult animals were preserved. Testes and epididymides were preserved in
Bouin’s solution, all other organs in 10% buffered formalin solution.

Postmortem examinations (offspring):
SACRIFICE/GROSS NECROPSY
Pups euthanized at PND 4 were carefully examined at least externally for gross abnormalities.

Statistics:
The statistical evaluation of appropriate data was performed with the statistical program package SPSS PC+4.0. The homogeneity of variance between groups was checked by Bartlett’s homogeneity of variance test. Where no significant heterogeneity was detected, a one-way analysis of variance (ANOVA) was carried out. If the obtained result was significant, Duncan Multiple Range test was used to access the significance of inter-group differences. Getting significant result at Bartlett’s test, the Kruskal-Wallis analysis of variance was used and the inter-group comparisons were performed using Mann-Whitney U-test. Chi2 test was performed if feasible.
Reproductive indices:
Calculation of Mating and Fertility Indice
- Male Mating Index: Number of males with confirmed mating/ Total Number of males cohabited x 100
- Female Mating Index: Number of sperm-positve females/ Total Number of females cohabited x 100
- Male Fertility Index: Number of males impregnating a female/ Total Number of males cohabited x 100
- Female Fertility Index: Number of pregnant females/ Number of sperm-positve females x 100
- Gestation Index: Number of females with live born pups/ Number of pregnant females x 100
Offspring viability indices:
Calculation of Pups’ Mortality and Sex Ratio Indices:
- Survival Index: Number of live pups (at designated time)/ Number of pups born x 100
- Pre-implantation mortality: (Number of Corpora lutea – Number of Implantations)/ Number of Corpora lutea x 100
- Intrauterine mortality: (Number of implantations – Number of liveborns)/ Number of implantations x 100
- Total mortality: Number of implantations/ (Number of implantation – Number of viable pups (d4)) x 100
- Sex ratio: (Number of pups examined – Number of males) / Number of pups examined x 100

Results and discussion

Results: P0 (first parental animals)

General toxicity (P0)

Clinical signs:
effects observed, treatment-related
Body weight and weight changes:
effects observed, treatment-related
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Gross pathological findings:
effects observed, treatment-related
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Other effects:
not examined

Reproductive function / performance (P0)

Reproductive function: estrous cycle:
effects observed, treatment-related
Reproductive function: sperm measures:
no effects observed
Reproductive performance:
effects observed, treatment-related

Details on results (P0)

CLINICAL SIGNS AND MORTALITY (PARENTAL ANIMALS):
Five female animals were found dead in the High dose group between Days 14 and 45. In addition one female treated at 600 mg/kg/day was found dead on Day 27. The death of this animal was accidental (gavage accident proved at necropsy).

Dark faeces were noted in all males and pregnant females treated at a dose level of 120 and 600 mg/kg/day. Limited use of hind-limbs was
observed in one male animal treated with 120 mg/kg/day.
In the surviving non-pregnant females treated at 600 mg/kg/day, decreased activity, dark faeces, hunched back, salivation, lethargy, piloerection and red liquid from the vulva were noted. The decreased activity, hunched back, piloerection and dark faeces were seen in all these High dose females.
In found dead animals, treatment-related clinical signs were observed including decreased activity, dark faeces, liquid faeces, hunched back, laboured respiration, lethargy, piloerection and red liquid from the mouth and vulva. Dark faeces were seen in all these animals.
In addition, occasionally clinical observations were noted, which were regarded as incidental finding and considered unrelated to treatment, missing
right testes; broken left incisor, and missing fur in the chin area.
No clinical signs were observed in animals treated at 25 mg/kg/day or in the control group.


BODY WEIGHT AND FOOD CONSUMPTION (PARENTAL ANIMALS)
Reduced body weight or body weight gain was observed in both sexes at 120 and 600 mg/kg/day; in males the terminal body weights were about 7% and 14% lower than control, in females the day 14 body weights were about 5% and 11% below controls at 120 and 600 mg/kg/day respectively,
although the effect became more pronounced in pregnancy. Food intake values were reduced in line with the body weight effects.
The Low dose group mean weights were unaffected by treatment.

REPRODUCTIVE FUNCTION: ESTROUS CYCLE (PARENTAL ANIMALS)
Toxicological significance effects were observed on the duration of the mating period in High dose animals. Successful coitus (sperm positive vaginal smears and/or vaginal plugs) occurred within up to 5 days of pairing (cohabitation), with the exception of High dose females 4503 and 4512, which did not have a sperm positive vaginal smear during the 14 days mating period. In addition, one High dose female animal no 4509 was found dead on the days 13 of the mating period, without successful coitus. Almost only dioestrus picture characterized the oestrus cycle of these animals.

REPRODUCTIVE FUNCTION: SPERM MEASURES (PARENTAL ANIMALS)
Histopathological evaluation of the male gonads as well as testicular interstitial cell structure, the spermatogenic cells representing different phases of the development and differentiation of the spermatozoa were similar in Control and High Dose males. There was no meaningful difference between incidence and severity of microscopic changes in the reproductive organs in High Dose males compared to Control males. Testicular observations had unilateral distribution in High Dose males. The incidence and severity were without a relationship to treatment and considered to be in the range of common background.

REPRODUCTIVE PERFORMANCE (PARENTAL ANIMALS)
The mating indices were 100% in the control, Low dose and Mid dose groups and 73% in the High dose group, while fertility indices were 100% in control, in Low and in Mid dose groups and 63% in High dose animals (due to 3/11 non-pregnant females). Gestation index was 92% in the control group (due to one animal no 1507 with stillborns, variation considered incidental), 100% in low dose group, 67% in Mid dose group (due to 4/12 animals with stillborns) and 0% in the High dose group.

ORGAN WEIGHTS (PARENTAL ANIMALS)
Males:
When compared to controls, statistically lower seminal vesicles weights were noted when measured as absolute values, and when adjusted for brain weights, in the High dose male animals (p<0.05). However, there was no histological evidence for any effects on the male reproductive system. Other statistical difference in male organ weights were ascribed to a consequence of body weight differences.

Females
When compared to controls, significantly lower vagina mean absolute weight was noted, about 47% below control in the High dose females (p<0.01).
This effect is probably related to ovary atrophy, since vagina weight is sensitive to changes in ovarian hormone production. The vagina weights of individual rats at this dose group show there was no influence of pregnancy on vaginal weight. There were a number of other statistical differences in relative organ weights, but at 120 and 600 mg/kg/day, weights of females at termination were about 18 % and 30 % below control, so the relative organ weight statistical differences were largely a consequence of body weight effects.

GROSS PATHOLOGY (PARENTAL ANIMALS)
In the six found dead High dose female animals, test item-related macroscopic changes were found in the variety of organs including black/grey discoloration/focus of the adrenal gland, brain, stomach, small intestines, caecum, colon, rectum, thymus, kidney, mesenteric lymph node and uterus. Pale discoloration of the liver was seen in 2 females. The small thymus present in one female was also regarded as test item-related and corresponded with similar change in terminal females from the High dose group.
Test item-related macroscopic findings were observed at dose levels of 120 and 600 mg/kg bw/day.
Four (4/6) females from the High Dose group were non-pregnant. A decrease or no corpora lutea and no implantation sites were seen in these animals at necropsy.
Small thymus in 4/6 High dose females and pale discoloration of the liver in 2/12 Mid dose and 1/6 High dose female rats, were present. Numerous organs were black/grey discoloured including the adrenal gland, brain, stomach, small intestines, caecum, colon, rectum, thymus, kidney, mesenteric lymph node, testis, ovary and/or uterus.


HISTOPATHOLOGY (PARENTAL ANIMALS)
Found dead animals
There were test item-related effects in the ovary, uterus, vagina, liver, kidney, thymus, mesenteric lymph node, stomach and caecum in 5 found dead females. One female was not histopathologically examined due to cannibalization of internal organs.

Minimal to moderate atrophy of the ovary, uterus and/or vagina in 3/5 females, moderate vacuolation of corpora lutea in the right ovary in 1/5 female, mild blue/black diffuse pigment deposits of the right ovary in 1/5 animal, were seen. In the liver, moderate hepatocellular necrosis and/or moderate diffuse vacuolation were found in 2/5 female rats. Mild bilateral necrosis of cortical tubules in the kidney associated with mild mixed cellular peritubular/perivascular infiltrate and moderate lymphoid atrophy of the thymus altered were seen in 1/5 found dead rat. Minimal or moderate accumulation of pigmented macrophages in the medulla of the mesenteric lymph node was noted in 3/5 animals. Mild mucosal erosion of the glandular stomach in 1/5 female and mild granular/crystalline foreign material in the lumen of caecum in 1/5 female rat, were also evaluated.
Other changes were incidental or agonal in nature and not related to treatment.


Scheduled necropsy
Test item-related findings were observed by light microscopy. At a dose level of 600 mg/kg bw/day, these changes were noted in the ovary, vagina, uterus , liver and mesenteric lymph node. In the females dosed at 120 mg/kg bw/day, the liver was only organ which was affected. These changes were characterized as atrophy of the ovary, uterus, vagina and/or thymus, blue/black pigment deposits in the ovary, accumulation of pigmented macrophages in mesenteric lymph node and hepatocellular vacuolation or necrosis in the liver. In the ovary, reduced number/size of follicles/corpora lutea, were observed. Occasionally, blue/black diffuse pigment deposits were present in the ovary. Low columnar luminal and glandular epithelium, reduced endometrium/myometrium were seen in the uterus. Attenuated epithelium comprising 2-3 cell layers was noted in the vagina. Lymphoid atrophy of thymus was accompanied with decreasing number of lymphocytes leading to decreased cell density, decreased compartment size (more pronounced in cortex), which also corresponded with organ weight change related brain weight. In the mesenteric lymph node, various degree of pigmented macrophages in the medulla was described. Vacuolation of hepatocytes contained predominantly large well-defined round vacuoles, with displaced nucleus to the periphery was one of the microscopic feature in the liver. Nuclei underwent lysis and increased eosinophilia of the cytoplasm, were typical for necrotic process in the liver. Mixed cell infiltrate could be also seen.
There was a consistent relationship to dose in the severity and incidence of these test item-related microscopic effects.
In the males, dose-related accumulation of pigmented macrophages was microscopically observed at dose levels 120 and 600 mg/kg bw/day in the mesenteric lymph node and corresponded with changes noted at necropsy.

Effect levels (P0)

open allclose all
Dose descriptor:
NOAEL
Effect level:
600 mg/kg bw/day (actual dose received)
Based on:
act. ingr.
Sex:
male
Basis for effect level:
other: Reproductive toxicity There were no effects identified on the male reproductive system.
Dose descriptor:
NOAEL
Effect level:
25 mg/kg bw/day (actual dose received)
Based on:
act. ingr.
Sex:
female
Basis for effect level:
other: Reproductive toxicity in parental females at 120 and 600 mg/kg/day was not considered secondary to systemic toxicity.
Dose descriptor:
NOAEL
Effect level:
25 mg/kg bw/day (actual dose received)
Based on:
act. ingr.
Sex:
male/female
Basis for effect level:
other: NOAEL for systemic effects of parental generation

Results: F1 generation

General toxicity (F1)

Clinical signs:
not examined
Mortality / viability:
mortality observed, treatment-related
Body weight and weight changes:
effects observed, treatment-related
Sexual maturation:
not examined
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
effects observed, treatment-related
Histopathological findings:
not examined

Details on results (F1)

VIABILITY (OFFSPRING)
There were no live born pups in the High dose group. From 19 examined pups, 16 pups were found dead and presented negative floating test and were not suckled and 3 were cannibalised on Day 0.
In the 120 mg/kg/day Mid dose group, 33/137 pups were possibly stillborn, 28/137 were found dead but born alive (presented positive floating test), 39 were cannibalized and 65 were not suckled. Of the pups which died PND0-PND4, there were more males than females.
In the Low dose, there was no effect on the number of live born pups (148/173). Lack of suckling was observed in 33 pups, one pup was pale and one was cold and cyanotic. The survival index of about 75% on PND4 is in below the normal control range.


CLINICAL SIGNS (OFFSPRING)
not examined

BODY WEIGHT (OFFSPRING)
Adverse effects considered related to Tellurium dioxide were observed with regard to offspring body weight on PND 0 in animals treated at 25, 120 and 600 mg/kg bw/day.
When compared to controls, the mean litter weights on PND 0, pups body weight evaluated on PND 0 for all pups or per litter, were lower in the F1 generation following administration of 25, 120 and 600 mg/kg/day daily by oral gavage, to the parental generation. These differences attained statistically significance in PND 0 body weight for all pup means (p<0.05 or p<0.01).
The pup body weight evaluated on PND 4 and the body weight gain was similar to the control in the Low and Mid dose animals. However, the total litter weights were considered to be below the normal control range in the Mid dose group on PND 0 and 4, and in the Low dose group at PND 0; these effects were ascribed to pup mortality rather than an effect on the weight of surviving pups.

The differences seen in the Low, Mid and High groups were considered to be a consequence of pup mortality, without an effect on the growth of survivors.

SEXUAL MATURATION (OFFSPRING)
not examined

ORGAN WEIGHTS (OFFSPRING)
not examined

GROSS PATHOLOGY (OFFSPRING)
Test item-related gross changes were observed in the cranium region and skin/subcutis in the High Dose group pups (two litters affected). In four found dead pups from the High Dose group, the absence of cranial region of the head with reduced brain size, covered by skin were noted. Whole body subcutaneous gelatinous material was recorded in 16 found dead pups.

HISTOPATHOLOGY (OFFSPRING)
not examined

OTHER FINDINGS (OFFSPRING)

Effect levels (F1)

Dose descriptor:
NOAEL
Generation:
F1
Effect level:
25 mg/kg bw/day (actual dose received)
Based on:
act. ingr.
Sex:
male/female
Basis for effect level:
other: pup mortality at 25 mg/kg/day
Remarks on result:
not determinable
Remarks:
no NOAEL identified

Overall reproductive toxicity

Reproductive effects observed:
not specified

Any other information on results incl. tables

No test item was identified in the control samples. The test item formulation appeared to be homogenous and had actual concentrations of 95.2-105.3% of the nominal concentrations, within the 100±15% acceptable range.

These results were considered suitable for the study purposes.

Applicant's summary and conclusion

Conclusions:
The No Effect Level for effects on the Parental generation was considered to be 25 mg/kg/day.
There was no No Effect Level for reproductive effects, due to pup mortality at 25 mg/kg/day.
There were no effects identified on the male reproductive system.
Executive summary:

In a screening study according to OECD guideline 421 Tellurium dioxide was administered by repeated oral gavage daily administration to Wistar rats at 0, 25, 120 and 600 mg/kg bw/day in 1% aqueous Methylcellulose. Males were dosed for 28 days (14 days pre-mating and 14 days mating/post- mating) and females were dosed for 14 days pre-mating, for up to 14 days mating period, through gestation and up to and including the day before necropsy, 4 days post-partum dosing.

 

At 600 mg/kg/day treatment related effects in females included mortality (5/12), clinical signs of decreased activity, hunched back, piloerection and dark faeces. In males and females at 120 and 600 mg/kg/day dark faeces was observed.

Reduced body weight or body weight gain was observed in both sexes at 120 and 600 mg/kg/day; in males the terminal body weights were about 7% and 14% lower than control, in females the day 14 body weights were about 5% and 11% below controls at 120 and 600 mg/kg/day respectively, although the effect became more pronounced in pregnancy. Food intake values were reduced in line with the body weight effects. The Low dose group mean weights were unaffected by treatment.

At 600 mg/kg/day there were adverse effects of treatment on the oestrus cycle, mating ability, fertility and gestation period. At 120 mg/kg/day the gestation period was prolonged. Higher intrauterine mortality, post-natal or total mortality were recorded at 120 and 600 mg/kg/day. Also at the Low dose, post-natal and total mortality was considered to be increased. There were no surviving pups at 600 mg/kg/day. Mortality data indicate a higher mortality in male pups than female pups in the 120 mg/kg/day group.

Examination of pups at 600 mg/kg/day, all were found dead, showed evidence of foetotoxic effects and possibly a developmental effect of the test item particularly on the cranial and brain development. The cause of death in pups at 25 and 120 mg/kg/day was not evident.

There were no evident effects on pup weight or weight gain in the surviving pups. Total litter weights at termination were reduced at 120 mg/kg/day this was considered to be a consequence of pup mortality.

At necropsy and histopathology, at 600 mg/kg/day there was a test item related atrophy of female reproductive tissues, which could account for the lack of mating and adverse embryo and foetal effects, with likely endocrine effects related to ovarian atrophy. There were signs of intestinal, hepatic and mesenteric lymph node toxicity, with some effects in kidney and thymus, there were pigment deposits in a number of affected tissues. The reproductive organ effects in females are not considered to be secondary effects of systemic toxicity. At 120 mg/kg/day females showed signs of hepatotoxicity but no apparent effects on reproductive organs. There were no effects identified on the male reproductive system.

 

The No Effect Level for effects on the Parental generation was considered to be 25 mg/kg/day.

There was no No Effect Level for reproductive effects, due to pup mortality at 25 mg/kg/day. Reproductive toxicity in parental females at 120 and 600 mg/kg/day was not considered secondary to systemic toxicity.