Registration Dossier

Toxicological information

Toxicity to reproduction

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Administrative data

Endpoint:
screening for reproductive / developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2013
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Cross-referenceopen allclose all
Reason / purpose for cross-reference:
reference to same study
Reference
Endpoint:
short-term repeated dose toxicity: oral
Remarks:
combined repeated dose and reproduction / developmental screening
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Study was conducted according to GLP and in compliance with agreed protocols, with no or minor deviations from standard test guidelines.
Reason / purpose for cross-reference:
reference to same study
Reason / purpose for cross-reference:
reference to same study
Qualifier:
according to guideline
Guideline:
OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)
Deviations:
no
Remarks:
No relevant deviation from the standard guideline
GLP compliance:
yes
Limit test:
no
Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals or test system and environmental conditions:
Details on animals:
Species/strain: Wistar rats, Crl: WI(Han) (Full Barrier)
Source: Charles River, 97633 Sulzfeld, Germany
Sex: male and female; the female animals were non-pregnant and nulliparous.
Age at the start of
the treatment period: males: 10-11 weeks old. females: 10-11 weeks old.
Body weight at the allocation of the animals to the experimental groups: males: 250 - 298 g (mean: 272.80 g, ± 20% = 218.24 – 327.36 g) - females: 162 - 199 g (mean: 183.13 g, ± 20% = 146.50 – 219.75 g)
The animals were derived from a controlled full-barrier maintained breeding system (SPF).

Housing and Feeding Conditions
- Full barrier in an air-conditioned room
- Temperature: 22 3°C
- Relative humidity: 55 10%
- Artificial light, sequence being 12 hours light, 12 hours dark
- Air change: 10 x / hour
- Free access to Altromin 1324 maintenance diet for rats and mice
- Free access to tap water, sulphur acidified to a pH of approximately 2.8 (drinking water, municipal residue control, microbiological controls at regular intervals)
- The animals were kept individually in IVC cages (except during the mating period when one female will be paired with one male), type III H, polysulphone cages on Altromin saw fibre bedding
- Adequate acclimatisation period (at least 5 days) under laboratory conditions


Route of administration:
oral: gavage
Vehicle:
CMC (carboxymethyl cellulose)
Remarks:
0.5 %
Details on oral exposure:
Preparation of the Animals
Prior to the start of the treatment period a detailed clinical observation outside the home cage was made. Before the first administration all animals used for the study were weighed and assigned to the experimental groups with achieving a most homogenous variation in body weight throughout the groups of males and females.

Experimental Groups and Doses
According to the results of the dose range finding study the following doses: 100 ; 300 and 1000 mg/kg bw were selected for the 3 dose groups (LD = low dose, MD = medium dose, HD = high dose) and 1 control group (C).

The test item and vehicle were administered at a single dose to the animals by oral gavage. The application volume for all groups was 5 mL/kg body weight. For each animal the individual dosing volume was calculated on the basis of the body weight.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Dose Formulation Analysis
Each dosing concentration was analyzed for nominal concentration by ICP - optical emission spectroscopy using a validated analytical procedure. Stability and homogeneity of the test item in the vehicle was analyzed for the LD, MD and HD dosing formulation.
Samples for the nominal concentration verification was taken in study week 1 (first week of pre mating period), 3 (first week of mating), 5 (gestation) and 7 (gestation/lactation) of control and all treatment groups.
Samples for homogeneity were taken from the top, middle and bottom of HD, MD, and LD preparation in study week 1 and 5.
Duration of treatment / exposure:
The animals were treated with the test item formulation or vehicle on 7 days per week for a period of 54 days, i.e. during 14 days of pre-mating and maximum 14 days of mating in both males and females, during the gestation period and up to post-natal day 3 in females.
Males were dosed after the mating period until the minimum total dosing period of 28 days was completed.
Frequency of treatment:
Once daily
Dose / conc.:
100 mg/kg bw/day (actual dose received)
Dose / conc.:
300 mg/kg bw/day (actual dose received)
Dose / conc.:
1 000 mg/kg bw/day (actual dose received)
No. of animals per sex per dose:
10
Control animals:
yes, concurrent vehicle
Details on study design:
According to the results of the dose range finding study and in consultation with the sponsor three selected doses were tested for the 3 dose groups (LD = low dose, MD = medium dose, HD = high dose) and 1 control group (C).
Observations and examinations performed and frequency:
Body Weight and Food Consumption
The body weight of all animals were recorded once before the assignment to the experimental groups. on the first day of administration and weekly during the treatment period as well as at the end of the study. During pregnancy, females were weighed on gestation days (GD) 0, 7, 14 and 20 and within 24 hours of parturition (day 0 post-partum) as well as day 4 post-partum along with pups.
Food consumption was measured weekly on the corresponding days of the body weight measurements after the beginning of the dose administration. Food consumption was not measured during the mating period in males and females and the post-mating period in males. The food consumption in males were measured only for two week during premating period and not during the postmating period as the number of days during the postmating period are not uniform in all males due to difference in mating.

Mating
Mating was performed in a ratio of 1:1 (male to female). The vaginal smears of the females were checked every morning after the start of the mating period to confirm the copulation. The day of the vaginal plug and/or sperm was considered as day 0 of gestation.
The cages were arranged in such a way that possible effects due to cage placement were minimised.
Copulation index, fertility index and delivery index was calculated for each group. The calculations were made using the formula as below,
Copulation Index (%) = (No. of rats copulated / No.of pairs) X 100
Fertility Index (%) = (No. of females pregant / No.of females copulated) X 100
Delivery Index (%) = (No. of dams with live newborns / No.of pregnant dams) X 100

Clinical Observations
General clinical observations were made once a day. Once before the first exposure, and once a week thereafter, detailed clinical observations were made in all animals outside the home cage in a standard arena. Clinical observations included spontaneous activity, lethargy, recumbent position, convulsions, tremors, apnoea, asphyxia, vocalization, diarrhea, changes in the skin and fur, eyes and mucous membranes (salivation, discharge), piloerection and pupil size, Changes in gait, posture, response to handling as well as the presence of clonic or tonic movements, stereotypes, difficult or prolonged parturition or bizarre behaviour were recorded.

Functional Observations

Multiple detailed behavioural observations were made in the week before the first treatment and during the last week of the treatment in 5 selected males and on lactation days in 5 selected females (only lactating females were evaluated) of control and treated groups outside the home cage using a functional observational battery of tests [11]:
Sensory reactivity to different modalities, grip strength and motor activity assessments and other behavioural observations as well as rearing supported and not supported, urination, defecation, startle/ auditory response, equilibrium reflex, positional passivity, visual placing, fore and hind limb grip strength, tail pinch response, toe pinch reflex, extensor thrust/limb tone, hind limb reflex, righting reflex on the ground, air righting reflex, pupil response, body temperature and ophthalmoscopy (anterior chamber of the eye and fundus of eye).

Litter Observations

The duration of the gestation was recorded and was calculated from day 0 of the pregnancy. Each litter was examined as soon as possible after delivery of the dam to establish the number and sex of pups, stillbirths, live births, runts and the presence of gross abnormalities.
The pre- and post- implantation losses were calculated using number of corpora lutea, number of implantation sites and number of live pups born on PND 0 for each dam. The formula used for the calculation are as follows,
Pre Implantation Loss (%) = (No. of corpora Lutea - No. of implantation site / No. of corpora Lutea) x 100
Post Implantation Loss (%) = (No. of implantation site – No. of live pups / No. of implantation site) x 100
Live pups were counted and sexed and weighed within 24 hours of parturition (day 0 post-partum) and on day 4 post-partum. Live pups were identified by tattooing. In addition to the observations of parent animals, any abnormal behaviour of the offspring was recorded.

Haematology
Haematological parameters from five selected males and females of each group were examined at the end of the treatment.
Blood from the abdominal aorta of the animals was collected in EDTA-coated tubes.
The following haematological parameters were examined:
Ihaematocrit value
haemoglobin content
red blood cell count
mean corpuscular volume (
mean corpuscular haemoglobin
mean corpuscular haemoglobin concentration
reticulocytes
platelet count
white blood cells
neutrophils
lymphocytes
monocytes
eosinophils
basophils

Blood Coagulation

Coagulation parameters from five selected males and females of each group were examined at the end of the treatment.
Blood from the abdominal aorta of the animals was collected in citrate- coated tubes.
The following coagulation parameters were examined:
prothrombin time
activated partial thromboplastin time

Clinical Biochemistry
Parameters of clinical biochemistry from five selected males and females of each group were examined at the end of the treatment.
Blood from the abdominal aorta of the animals was collected in serum separator tubes.
The following parameters of clinical biochemistry were examined:
alanine aminotransferase
aspartate-aminotransferase
alkaline phosphatase
creatinine
total protein
albumin
urea
total bile acids
total cholesterol
glucose
sodium
potassium
Calcium
Phosphorus

Urinalysis

A urinalysis was performed with samples collected from 5 selected males of each group at necropsy. Additionally, urine colour/ appearance were recorded.
The following parameters were measured using qualitative indicators (Heiland Urine Stripes URI 10SL).
specific gravity
nitrite
ph-value
protein
glucose
ketone bodies
urobilinogen
bilirubin
blood
leukocytes

Sperm analysis
At necropsy (one day after the last administration) one epididymis and one testis from males of each group were separated and used for evaluation of sperm parameters. Epididymal sperm motility was evaluated in all male animals using Hamilton Thorn Sperm Analyser. The testicular sperm count could not be measured at the end of the study as the testes stored at -80oC were inadvertently taken out of the freezer before the scheduled measurement.

Sacrifice and pathology:
Gross necropsy
All male animals were sacrificed after the completion of the mating period (minimum dosing period: 28 days) on study day 29 or 30, while female animals were sacrificed on post-natal day 4 (maximum dosing period: 54 days) using an anaesthesia was used.
Females showing no sign of pregnancy was sacrificed on day 26 after the last day of the mating period.
All animals were subjected to a detailed gross necropsy which includes careful examination of the external surface of the body, all orifices and the cranial, thoracic and abdominal cavities and their contents.

Organ Weights
The wet weight of the organsof 5 males and 5 females selected from each group was recorded as soon as possible. Paired organs were weighed separately. In addition reproductive organs of all animals were weighed. The enclosed organs were weighed at necropsy:
liver
uterus with cervix
kidneys
thymus
adrenals
thyroid/parathyroid glands
testes
spleen
epididymides
brain
prostate. seminal vesicles and coagulating glands
pituitary gland
ovaries

The following tissues of the same selected animals from each group were preserved in 10% neutral buffered formalin except eyes, testes and epididymides that were fixed in Modified Davidson’s Fixative for approximately 24 hours before they were transferred to 10% neutral buffered formalin:
brain (cerebrum, cerebellum and pons)
ovaries (females)
spinal cord
uterus with cervix (females)
liver
vagina (females)
kidneys
testes (males)
adrenal glands
epididymides (males)
stomach
prostate and seminal vesicles with coagulating glands as a whole (males)
small and large intestines (including Peyer´s patches)
urinary bladder
thymus
lymphnodes (mesentric and axillary)
Thyroid
peripheral nerve (e.g. sciatic nerve) with skeletal muscle
spleen
bone with bone marrow (sternum)
lung and trachea
pituitary gland
mammary glands
oesophagus
heart
gross lesions

Histopathology
All organs and tissues listed in Table 7 were evaluated from five selected males and females of the control and high dose group:
Reproductive organsand macroscopic changes were evaluated in all study animals. For the testes, a detailed qualitative examination was made.

Statistics:
Evaluation of Results and Statistical Analysis
The findings of this study were evaluated in terms of the observed effects, the necropsy and the microscopic findings.
Parameters like body weight gain and food consumption were calculated for each animal as the difference in weight measured from one week to the next. Mean body weights are also presented as figures. The relative organ weights were calculated in relation to the body weight (measured at necropsy) and are presented as percentage.
A statistical assessment of the results of the body weight. food consumption, parameters of haematology, blood coagulation and clinical biochemistry and absolute and relative organ weights were performed for each gender by comparing values of dosed with control animals of the main groups using a one-way ANOVA and a post-hoc Dunnett Test. These statistics were performed with GraphPad Prism 5.01 software (p<0.05 was considered as statistically significant).
Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
There were no clinical signs recorded in male and female animals of treated groups that could be directly related to treatment. However, there were few clinical signs namely moving the bedding, nasal discharge (dark or reddish), salivation and piloerection seen occasionally and transiently during the study period in MD or HD group animals. These findings were considered to be due to local effect but not the systemic effect of the test item. These findings were considered not likely to be adverse. In addition, there was alopecia (on hind limb, forelimb, thorax and abdominal region) of few isolated animals of MD or HD groups, which was assumed to be incidental in origin.
During the weekly detailed clinical observation, no significant changes or differences between the groups were found.
Mortality:
no mortality observed
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
In both males and females, no treatment related changes were noted for body weight and body weight change during the study period. Statistically there were significant increase in body weight change in female HD group during 2nd week of premating period when compared to control. In addition, there was lower mean body weight gain noted between days 1-7 of premating when compared to control without attaining the statistical significance. But, this increase or decrease in weight gain did not correlate with food intake during the same period. Hence, the changes were not considered likely to be adverse. There was decrease in body weight gain noted in female MD and HD groups during lactation period when compared to control. This decrease had no statistical significance and was not likely to be adverse.
Food consumption and compound intake (if feeding study):
no effects observed
Description (incidence and severity):
In both males and females, no treatment related changes were noted for treated group when compared to corresponding control. The statistical evaluation of the data revealed no significant changes in food intake in treated group animals when compared to control.
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
no effects observed
Ophthalmological findings:
no effects observed
Description (incidence and severity):
There were no ophthalmoscopic findings in any of the animals of this study.
Haematological findings:
no effects observed
Description (incidence and severity):
No treatment related changes were noted for haematology and blood coagulation parameters measured at the end of treatment period in male and female animals. There were no statistically significant changes noted for haematological and coagulation parameters between the treated and control groups.
Clinical biochemistry findings:
effects observed, non-treatment-related
Description (incidence and severity):
There were no treatment related changes considered for measured clinical biochemistry parameters of male and female treated groups when compared to corresponding control. However, there was statistically significant increase noted for mean potassium value in male MD group. In the absence of dose response pattern no relevance to treatment was considered.
Urinalysis findings:
no effects observed
Description (incidence and severity):
The urinalysis performed in male animals revealed no considerable changes in treated groups when compared to the control.
Behaviour (functional findings):
no effects observed
Description (incidence and severity):
No relevant effects of treatment were observed in any of the parameters of the functional observation battery before and at the end of the treatment period. There were no biologically relevant differences in body temperature between the groups.
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
There were no changes considered to be related to treatment noted for organ weight in both males and females when compared to corresponding control. However, there was statistically significant increase in relative weight of left kidney weight in male treated (LD, MD and HD) groups, but not total kidney weight. This change in left kidney weight, in the absence of histological changes was not considered to have toxicological relevance.
Gross pathological findings:
effects observed, non-treatment-related
Description (incidence and severity):
At terminal sacrifice, macroscopic organ findings noted were few, and none of them was considered to be test item related.
Yellow spot(s) at the epididymis were observed without dose relationship in 2/10 control males, 1/10 male of LD group, 3/10 males of MD group and 2/10 males of HD group. They were not considered treatment related as histologically they were confirmed to represent spermatic granuloma, a finding known to occur spontaneously in untreated rats of this strain and age.
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
Reproductive organs
No test item-related effects were noted on male and female reproductive organs in any of the treatment groups.
Histopathological findings noted in male reproductive organs were few and considered to be spontaneous in nature and unrelated to the test item, comprising multifocal atrophic tubules in the testis of each one male treated at 300 and 1000 mg/kg/day and subsequent epididymal changes in the same animals.
Reproductive organs of most study females showed histomorphological evidence of precedent pregnancy in the uterus. The number of large corpora lutea in the ovary was not essentially different between the groups.
Non pregnant animals showed physiological sexual cycling, and their unsuccessful mating was considered unrelated to the treatment.
Other organs
No test item-related histopathological findings were noted in the other organs evaluated in randomized males and females of the control and high dose group.
Histopathological changes seen at terminal sacrifice were considered to be incidental in origin and/or within the range of expected changes for rats of this age and strain kept under laboratory conditions.
Key result
Dose descriptor:
NOAEL
Effect level:
>= 1 000 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: No adverse effects observed at the highest dose tested.
Key result
Critical effects observed:
no
Conclusions:
Repeated dose administration of yttrium oxide to the male (minimum 28 days) and female (maximum 54 days) Wistar rats at dosages of 100, 300 and 1000 mg/kg body weight revealed neither mortalities nor findings of toxicological relevance in male and female animals. There were no toxicologically relevant findings noted for reproductive and developmental parameters.

Based on the data generated from this “Combined Repeated Dose Oral Toxicity Study with the Reproduction/ Developmental Toxicity Screening Test with Yttrium Oxide, the no observed adverse effect level (NOAEL) for systemic and reproductive and developmental toxicity is considered to be 1000 mg/kg body weight.

No classification for repeat-dose toxicity is warranted based on the absence of toxicologically relevant effects in this study, according to the criteria of Annex VI Directive 67/748/EEC or the 1272/2008 regulation -CLP).
Executive summary:

There was no mortality noted in treated (at 100, 300 and 1000 mg/kg bw/day) and control groups during the study period (at least 28 days for males and 54 days for females).

During the weekly detailed clinical observation, no significant changes or differences between the groups were found.

There were no ophthalmoscopic findings in any of the animals of this study.

Weight increase or decrease had no statistical significance and was not likely to be adverse.

The statistical evaluation of food consumption revealed no significant changes in food intake in treated group animals when compared to control.

There were no statistically significant changes noted nor for haematological and coagulation parameters nor for clinical biochemistry parameters of male and female treated groups when compared to corresponding control.

The urinalysis performed in male animals revealed no considerable changes in treated groups when compared to the control.

There were no changes considered to be related to treatment noted for organ weight in both males and females when compared to corresponding control. However, there was statistically significant increase in relative weight of left kidney weight in male treated (LD, MD and HD) groups, but not total kidney weight. This change in left kidney weight, in the absence of histological changes was not considered to have toxicological relevance.

At terminal sacrifice, macroscopic organ findings noted were few, and none of them was considered to be test item related.

No test item-related histopathological findings were noted in the other organs evaluated in randomized males and females of the control and high dose group.

Reason / purpose for cross-reference:
reference to same study
Reference
Endpoint:
developmental toxicity
Remarks:
screening study
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2013
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Reason / purpose for cross-reference:
reference to same study
Reason / purpose for cross-reference:
reference to same study
Qualifier:
according to guideline
Guideline:
other: OECD422
Deviations:
no
GLP compliance:
yes
Limit test:
no
Species:
rat
Strain:
Wistar
Details on test animals or test system and environmental conditions:
Details on animals:
Species/strain: Wistar rats, Crl: WI(Han) (Full Barrier)
Source: Charles River, 97633 Sulzfeld, Germany
Sex: male and female; the female animals were non-pregnant and nulliparous.
Age at the start of
the treatment period: males: 10-11 weeks old. females: 10-11 weeks old.
Body weight at the allocation of the animals to the experimental groups: males: 250 - 298 g (mean: 272.80 g, ± 20% = 218.24 – 327.36 g) - females: 162 - 199 g (mean: 183.13 g, ± 20% = 146.50 – 219.75 g)
The animals were derived from a controlled full-barrier maintained breeding system (SPF).

Housing and Feeding Conditions
- Full barrier in an air-conditioned room
- Temperature: 22 3°C
- Relative humidity: 55 10%
- Artificial light, sequence being 12 hours light, 12 hours dark
- Air change: 10 x / hour
- Free access to Altromin 1324 maintenance diet for rats and mice
- Free access to tap water, sulphur acidified to a pH of approximately 2.8 (drinking water, municipal residue control, microbiological controls at regular intervals)
- The animals were kept individually in IVC cages (except during the mating period when one female will be paired with one male), type III H, polysulphone cages on Altromin saw fibre bedding
- Adequate acclimatisation period (at least 5 days) under laboratory conditions

Route of administration:
oral: gavage
Vehicle:
CMC (carboxymethyl cellulose)
Remarks:
0.5%
Details on exposure:
Prior to the start of the treatment period a detailed clinical observation outside the home cage was made. Before the first administration all animals used for the study were weighed and assigned to the experimental groups with achieving a most homogenous variation in body weight throughout the groups of males and females.

Experimental Groups and Doses
According to the results of the dose range finding study the following doses: 100 ; 300 and 1000 mg/kg bw were selected for the 3 dose groups (LD = low dose, MD = medium dose, HD = high dose) and 1 control group (C).

The test item and vehicle were administered at a single dose to the animals by oral gavage. The application volume for all groups was 5 mL/kg body weight. For each animal the individual dosing volume was calculated on the basis of the body weight.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Dose Formulation Analysis
Each dosing concentration was analyzed for nominal concentration by ICP - optical emission spectroscopy using a validated analytical procedure. Stability and homogeneity of the test item in the vehicle was analyzed for the LD, MD and HD dosing formulation.
Samples for the nominal concentration verification was taken in study week 1 (first week of pre mating period), 3 (first week of mating), 5 (gestation) and 7 (gestation/lactation) of control and all treatment groups.
Samples for homogeneity were taken from the top, middle and bottom of HD, MD, and LD preparation in study week 1 and 5.
Details on mating procedure:
Mating was performed in a ratio of 1:1 (male to female). The vaginal smears of the females were checked every morning after the start of the mating period to confirm the copulation. The day of the vaginal plug and/or sperm was considered as day 0 of gestation.
The cages were arranged in such a way that possible effects due to cage placement were minimised.
Duration of treatment / exposure:
7d/wk for a period of 54 days, i.e. during 14 days of pre-mating and maximum 14 days of mating in both males and females, during the gestation period and up to post-natal day 3 in females. Males were dosed after the mating period until the minimum total dosing period of 28 days was completed.
Frequency of treatment:
Once daily
Duration of test:
Males minimum 28 days and females maximum of 54 days
Dose / conc.:
100 mg/kg bw/day (actual dose received)
Dose / conc.:
300 mg/kg bw/day (actual dose received)
Dose / conc.:
1 000 mg/kg bw/day (actual dose received)
No. of animals per sex per dose:
10
Control animals:
yes, concurrent vehicle
Details on study design:
According to the results of the dose range finding study and in consultation with the sponsor three selected doses were tested for the 3 dose groups (LD = low dose, MD = medium dose, HD = high dose) and 1 control group (C).
Maternal examinations:
Body Weight and Food Consumption
The body weight of all animals were recorded once before the assignment to the experimental groups. on the first day of administration and weekly during the treatment period as well as at the end of the study. During pregnancy, females were weighed on gestation days (GD) 0, 7, 14 and 20 and within 24 hours of parturition (day 0 post-partum) as well as day 4 post-partum along with pups.
Food consumption was measured weekly on the corresponding days of the body weight measurements after the beginning of the dose administration. Food consumption was not measured during the mating period in males and females and the post-mating period in males. The food consumption in males were measured only for two week during premating period and not during the postmating period as the number of days during the postmating period are not uniform in all males due to difference in mating.
Gross necropsy
All male animals were sacrificed after the completion of the mating period (minimum dosing period: 28 days) on study day 29 or 30, while female animals were sacrificed on post-natal day 4 (maximum dosing period: 54 days) using an anaesthesia was used. Pups were killed on PND 4 by using high dose of sodium pentobarbitol.
Pups sacrificed on day 4 post-partum were carefully examined externally for gross abnormalities.

Females showing no sign of pregnancy was sacrificed on day 26 after the last day of the mating period.
All animals were subjected to a detailed gross necropsy which includes careful examination of the external surface of the body, all orifices and the cranial, thoracic and abdominal cavities and their contents.
The number of implantation sites and corpora lutea was recorded for each parental female at necropsy.

Organ Weights
The wet weight of the organs of 5 males and 5 females selected from each group was recorded as soon as possible. Paired organs were weighed separately. In addition reproductive organs of all animals were weighed. The enclosed organs were weighed at necropsy:
liver
uterus with cervix
kidneys
thymus
adrenals
thyroid/parathyroid glands
testes
spleen
epididymides
brain
prostate. seminal vesicles and coagulating glands
pituitary gland
ovaries

The following tissues of the same selected animals from each group were preserved in 10% neutral buffered formalin except eyes, testes and epididymides that were fixed in Modified Davidson’s Fixative for approximately 24 hours before they were transferred to 10% neutral buffered formalin:
brain (cerebrum, cerebellum and pons)
ovaries (females)
spinal cord
uterus with cervix (females)
liver
vagina (females)
kidneys
testes (males)
adrenal glands
epididymides (males)
stomach
prostate and seminal vesicles with coagulating glands as a whole (males)
small and large intestines (including Peyer´s patches)
urinary bladder
thymus
lymphnodes (mesentric and axillary)
Thyroid
peripheral nerve (e.g. sciatic nerve) with skeletal muscle
spleen
bone with bone marrow (sternum)
lung and trachea
pituitary gland
mammary glands
oesophagus
heart
gross lesions

Histopathology
All organs and tissues were evaluated from five selected males and females of the control and high dose group:
Reproductive organs (ovary, uterus, cervix, vagina, testis, epididymis, prostate gland, seminal vesicle and coagulating gland) and macroscopic changes were evaluated in all study animals. For the testes, a detailed qualitative examination was made; taking into account the tubular stages of the spermatogenic cycle for the evaluation of additional hematoxylin-PAS (Periodic Acid Schiff) stained slides.
Fetal examinations:
The duration of the gestation was recorded and was calculated from day 0 of the pregnancy. Each litter was examined as soon as possible after delivery of the dam to establish the number and sex of pups, stillbirths, live births, runts and the presence of gross abnormalities.
The pre- and post- implantation losses were calculated using number of corpora lutea, number of implantation sites and number of live pups born on PND 0 for each dam. The formula used for the calculation are as follows,
Pre Implantation Loss (%) = (No. of corpora Lutea - No. of implantation site / No. of corpora Lutea) x 100
Post Implantation Loss (%) = (No. of implantation site – No. of live pups / No. of implantation site) x 100
Live pups were counted and sexed and weighed within 24 hours of parturition (day 0 post-partum) and on day 4 post-partum. Live pups were identified by tattooing. In addition to the observations of parent animals, any abnormal behaviour of the offspring was recorded.
Pups were killed on PND 4 by using high dose of sodium pentobarbitol.
Pups sacrificed on day 4 post-partum were carefully examined externally for gross abnormalities.
Statistics:
Evaluation of Results and Statistical Analysis
The findings of this study were evaluated in terms of the observed effects, the necropsy and the microscopic findings.
Parameters like body weight gain and food consumption were calculated for each animal as the difference in weight measured from one week to the next. Mean body weights are also presented as figures. The relative organ weights were calculated in relation to the body weight (measured at necropsy) and are presented as percentage.
A statistical assessment of the results of the body weight. food consumption, parameters of haematology, blood coagulation and clinical biochemistry and absolute and relative organ weights were performed for each gender by comparing values of dosed with control animals of the main groups using a one-way ANOVA and a post-hoc Dunnett Test. These statistics were performed with GraphPad Prism 5.01 software (p<0.05 was considered as statistically significant).
Indices:
Copulation index, fertility index and delivery index was calculated for each group. The calculations were made using the formula as below,
Copulation Index (%) = (No. of rats copulated / No.of pairs) X 100
Fertility Index (%) = (No. of females pregant / No.of females copulated) X 100
Delivery Index (%) = (No. of dams with live newborns / No.of pregnant dams) X 100
Clinical signs:
effects observed, non-treatment-related
Description (incidence and severity):
There were no clinical signs recorded in male and female animals of treated groups that could be directly related to treatment. However, there were few clinical signs namely moving the bedding, nasal discharge (dark or reddish), salivation and piloerection seen occasionally and transiently during the study period in MD or HD group animals. These findings were considered to be due to local effect but not the systemic effect of the test item. These findings were considered not likely to be adverse. In addition, there was alopecia (on hind limb, forelimb, thorax and abdominal region) of few isolated animals of MD or HD groups, which was assumed to be incidental in origin.
During the weekly detailed clinical observation, no significant changes or differences between the groups were found.
Mortality:
no mortality observed
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
In both males and females, no treatment related changes were noted for body weight and body weight change during the study period. Statistically there were significant increase in body weight change in female HD group during 2nd week of premating period when compared to control. In addition, there was lower mean body weight gain noted between days 1-7 of premating when compared to control without attaining the statistical significance. But, this increase or decrease in weight gain did not correlate with food intake during the same period. Hence, the changes were not considered likely to be adverse. There was decrease in body weight gain noted in female MD and HD groups during lactation period when compared to control. This decrease had no statistical significance and was not likely to be adverse.
Ophthalmological findings:
no effects observed
Description (incidence and severity):
There were no ophthalmoscopic findings in any of the animals of this study.
Behaviour (functional findings):
no effects observed
Description (incidence and severity):
No relevant effects of treatment were observed in any of the parameters of the functional observation battery before and at the end of the treatment period. There were no biologically relevant differences in body temperature between the groups.
Pre- and post-implantation loss:
no effects observed
Description (incidence and severity):
No treatment related changes were noted for number of corpora lutea, number of implantation sites, number of live pups born on PND 0 and percentage of pre and post implantation loss in treated groups when compared to control. The statistical evaluation of data revealed no significant differences between the values of treated and control groups.
Key result
Dose descriptor:
NOAEL
Effect level:
>= 1 000 mg/kg bw/day (nominal)
Based on:
test mat.
Basis for effect level:
other: maternal toxicity
Key result
Abnormalities:
no effects observed
Description (incidence and severity):
No treatment related changes were noted for survival of the pups from PND 0 to PND 4 in treated groups when compared to control. However, there was 1 pup each in Control (Pup no. 3; animal 50), LD (Pup no. 12, animal 58) and MD (Pup no. 9, animal 70) groups found dead or missing between PND 0 and 4. The missing pup was assumed to be cannibalized by dam, which was considered to be incidental. No treatment related changes were considered for viability index (%).
External malformations:
effects observed, non-treatment-related
Description (incidence and severity):
No treatment related gross external findings were observed in any of the treated groups. However, there were few isolated findings noted in pups namely dry skin, dark spot on/ or dark abdomen, dark snout in control group; dark spot on forelimb and thoracic back, dark snout in LD group; dry skin in MD and HD groups. These findings were considered to be incidental in origin.
Key result
Dose descriptor:
NOAEL
Effect level:
>= 1 000 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: no adverse effects seen at highest dose tested
Key result
Abnormalities:
no effects observed
Key result
Developmental effects observed:
no
Conclusions:
Repeated dose administration of Yttrium Oxide to male (minimum 28 days) and female (maximum 54 days) Wistar rats following OECD guideline 422 at dosages of 100, 300 and 1000 mg/kg body weight revealed neither mortalities nor findings of toxicological relevance in male and female animals. There were also no toxicologically relevant findings noted for reproductive and developmental parameters.

Based on these data, the no observed adverse effect level (NOAEL) for systemic and reproductive and developmental toxicity is considered to be 1000 mg/kg body weight/day. The NOEL for reproductive performance (mating and fertility) was considered to be 1000 mg/kg/day and the NOEL for toxic effects on progeny was 1000 mg/kg/day.
Executive summary:

A repeated dose study was performed according to OECD guideline 422 with Yttrium Oxide. Male (minimum 28 days) and female (maximum 54 days) Wistar rats were dosed at dosages of 100, 300 and 1000 mg/kg body weight/day. There were no changes considered to be related to treatment noted for organ weight in both males and females when compared to corresponding control. However, there was statistically significant increase in relative weight of left kidney weight in male treated (LD, MD and HD) groups, but not total kidney weight. This change in left kidney weight, in the absence of histological changes was not considered to have toxicological relevance.

No test item-related effects were noted on male and female reproductive organs in any of the treatment groups. The statistical analysis of epididymal sperm motility indicated no statistical significant difference between the treated and control groups. No treatment related changes were noted for the precoital interval and duration of gestation in treated groups when compared to control. All pregnancies resulted in normal births.

There were no treatment related changes noted for copulation index (%), fertility index (%), delivery index (%) and viability index (%) in treated groups when compared to corresponding control group.

Successful mating resulted 100% pregnancy rates in C and HD groups and 90% pregnancy rates in LD and MD groups. Histologically, they showed physiological sexual cycling, and their unsuccessful mating was considered unrelated to the treatment. The statistical evaluation of pre and post-natal data revealed no significant differences between the values of treated and control groups. No treatment related no significant changes were noted for survival of the pups and for viability index (%).

Based on these data, the no observed adverse effect level (NOAEL) for systemic and reproductive and developmental toxicity is considered to be 1000 mg/kg body weight/day. The NOEL for reproductive performance (mating and fertility) was considered to be 1000 mg/kg/day and the NOEL for toxic effects on progeny was 1000 mg/kg/day.

Data source

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

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)
Deviations:
no
GLP compliance:
yes
Limit test:
no

Test material

Constituent 1
Chemical structure
Reference substance name:
Yttrium oxide
EC Number:
215-233-5
EC Name:
Yttrium oxide
Cas Number:
1314-36-9
Molecular formula:
O3Y2
IUPAC Name:
Yttrium (III) oxide
Test material form:
solid: particulate/powder
Details on test material:
Chemical Name: Yttrium Oxide
Physical state: Fine Powder (at room temperature)
Colour: white

Test animals

Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals or test system and environmental conditions:
Details on animals:
Species/strain: Wistar rats, Crl: WI(Han) (Full Barrier)
Source: Charles River, 97633 Sulzfeld, Germany
Sex: male and female; the female animals were non-pregnant and nulliparous.
Age at the start of
the treatment period: males: 10-11 weeks old. females: 10-11 weeks old.
Body weight at the allocation of the animals to the experimental groups: males: 250 - 298 g (mean: 272.80 g, ± 20% = 218.24 – 327.36 g) - females: 162 - 199 g (mean: 183.13 g, ± 20% = 146.50 – 219.75 g)
The animals were derived from a controlled full-barrier maintained breeding system (SPF).

Housing and Feeding Conditions
- Full barrier in an air-conditioned room
- Temperature: 22 3°C
- Relative humidity: 55 10%
- Artificial light, sequence being 12 hours light, 12 hours dark
- Air change: 10 x / hour
- Free access to Altromin 1324 maintenance diet for rats and mice
- Free access to tap water, sulphur acidified to a pH of approximately 2.8 (drinking water, municipal residue control, microbiological controls at regular intervals)
- The animals were kept individually in IVC cages (except during the mating period when one female will be paired with one male), type III H, polysulphone cages on Altromin saw fibre bedding
- Adequate acclimatisation period (at least 5 days) under laboratory conditions

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
CMC (carboxymethyl cellulose)
Remarks:
0.5 %
Details on exposure:
Prior to the start of the treatment period a detailed clinical observation outside the home cage was made. Before the first administration all animals used for the study were weighed and assigned to the experimental groups with achieving a most homogenous variation in body weight throughout the groups of males and females.

Experimental Groups and Doses
According to the results of the dose range finding study the following doses: 100 ; 300 and 1000 mg/kg bw were selected for the 3 dose groups (LD = low dose, MD = medium dose, HD = high dose) and 1 control group (C).

The test item and vehicle were administered at a single dose to the animals by oral gavage. The application volume for all groups was 5 mL/kg body weight. For each animal the individual dosing volume was calculated on the basis of the body weight.
Details on mating procedure:
Mating was performed in a ratio of 1:1 (male to female). The vaginal smears of the females were checked every morning after the start of the mating period to confirm the copulation. The day of the vaginal plug and/or sperm was considered as day 0 of gestation.
The cages were arranged in such a way that possible effects due to cage placement were minimised.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Dose Formulation Analysis
Each dosing concentration was analyzed for nominal concentration by ICP - optical emission spectroscopy using a validated analytical procedure. Stability and homogeneity of the test item in the vehicle was analyzed for the LD, MD and HD dosing formulation.
Samples for the nominal concentration verification was taken in study week 1 (first week of pre mating period), 3 (first week of mating), 5 (gestation) and 7 (gestation/lactation) of control and all treatment groups.
Samples for homogeneity were taken from the top, middle and bottom of HD, MD, and LD preparation in study week 1 and 5.
Duration of treatment / exposure:
The animals were treated with the test item formulation or vehicle on 7 days per week for a period of 54 days, i.e. during 14 days of pre-mating and maximum 14 days of mating in both males and females, during the gestation period and up to post-natal day 3 in females.
Males were dosed after the mating period until the minimum total dosing period of 28 days was completed.
Frequency of treatment:
Once daily
Doses / concentrationsopen allclose all
Dose / conc.:
1 000 mg/kg bw/day (actual dose received)
Dose / conc.:
300 mg/kg bw/day (actual dose received)
Dose / conc.:
100 mg/kg bw/day (actual dose received)
No. of animals per sex per dose:
10 animals per sex per dose.

Control animals:
yes, concurrent vehicle
Details on study design:
According to the results of the dose range finding study and in consultation with the sponsor three selected doses were tested for the 3 dose groups (LD = low dose, MD = medium dose, HD = high dose) and 1 control group (C).

Examinations

Parental animals: Observations and examinations:
Body Weight and Food Consumption
The body weight of all animals were recorded once before the assignment to the experimental groups. on the first day of administration and weekly during the treatment period as well as at the end of the study. During pregnancy, females were weighed on gestation days (GD) 0, 7, 14 and 20 and within 24 hours of parturition (day 0 post-partum) as well as day 4 post-partum along with pups.
Food consumption was measured weekly on the corresponding days of the body weight measurements after the beginning of the dose administration. Food consumption was not measured during the mating period in males and females and the post-mating period in males. The food consumption in males were measured only for two week during premating period and not during the postmating period as the number of days during the postmating period are not uniform in all males due to difference in mating.
Oestrous cyclicity (parental animals):
The vaginal smears of the females were checked every morning after the start of the mating period to confirm the copulation. The day of the vaginal plug and/or sperm was considered as day 0 of gestation.
Sperm parameters (parental animals):
At necropsy (one day after the last administration) one epididymis and one testis from males of each group were separated and used for evaluation of sperm parameters. Epididymal sperm motility was evaluated in all male animals using Sperm Analyser. The testicular sperm count could not be measured at the end of the study as the testes stored at -80oC were inadvertently taken out of the freezer before the scheduled measurement.
Litter observations:
The duration of the gestation was recorded and was calculated from day 0 of the pregnancy. Each litter was examined as soon as possible after delivery of the dam to establish the number and sex of pups, stillbirths, live births, runts and the presence of gross abnormalities.
The pre- and post- implantation losses were calculated using number of corpora lutea, number of implantation sites and number of live pups born on PND 0 for each dam. The formula used for the calculation are as follows,
Pre Implantation Loss (%) = (No. of corpora Lutea - No. of implantation site / No. of corpora Lutea) x 100
Post Implantation Loss (%) = (No. of implantation site – No. of live pups / No. of implantation site) x 100
Live pups were counted and sexed and weighed within 24 hours of parturition (day 0 post-partum) and on day 4 post-partum. Live pups were identified by tattooing. In addition to the observations of parent animals, any abnormal behaviour of the offspring was recorded.
Postmortem examinations (parental animals):
Gross necropsy
All male animals were sacrificed after the completion of the mating period (minimum dosing period: 28 days) on study day 29 or 30, while female animals were sacrificed on post-natal day 4 (maximum dosing period: 54 days) using an anaesthesia was used. Pups were killed on PND 4 by using high dose of sodium pentobarbitol.
Pups sacrificed on day 4 post-partum were carefully examined externally for gross abnormalities.

Females showing no sign of pregnancy was sacrificed on day 26 after the last day of the mating period.
All animals were subjected to a detailed gross necropsy which includes careful examination of the external surface of the body, all orifices and the cranial, thoracic and abdominal cavities and their contents.
The number of implantation sites and corpora lutea was recorded for each parental female at necropsy.

Organ Weights
The wet weight of the organs of 5 males and 5 females selected from each group was recorded as soon as possible. Paired organs were weighed separately. In addition reproductive organs of all animals were weighed. The enclosed organs were weighed at necropsy:
liver
uterus with cervix
kidneys
thymus
adrenals
thyroid/parathyroid glands
testes
spleen
epididymides
brain
prostate. seminal vesicles and coagulating glands
pituitary gland
ovaries

The following tissues of the same selected animals from each group were preserved in 10% neutral buffered formalin except eyes, testes and epididymides that were fixed in Modified Davidson’s Fixative for approximately 24 hours before they were transferred to 10% neutral buffered formalin:
brain (cerebrum, cerebellum and pons)
ovaries (females)
spinal cord
uterus with cervix (females)
liver
vagina (females)
kidneys
testes (males)
adrenal glands
epididymides (males)
stomach
prostate and seminal vesicles with coagulating glands as a whole (males)
small and large intestines (including Peyer´s patches)
urinary bladder
thymus
lymphnodes (mesentric and axillary)
Thyroid
peripheral nerve (e.g. sciatic nerve) with skeletal muscle
spleen
bone with bone marrow (sternum)
lung and trachea
pituitary gland
mammary glands
oesophagus
heart
gross lesions

Histopathology
All organs and tissues were evaluated from five selected males and females of the control and high dose group:
Reproductive organs (ovary, uterus, cervix, vagina, testis, epididymis, prostate gland, seminal vesicle and coagulating gland) and macroscopic changes were evaluated in all study animals. For the testes, a detailed qualitative examination was made; taking into account the tubular stages of the spermatogenic cycle for the evaluation of additional hematoxylin-PAS (Periodic Acid Schiff) stained slides.

Postmortem examinations (offspring):
Pups were killed on PND 4 by using high dose of sodium pentobarbitol.
Pups sacrificed on day 4 post-partum were carefully examined externally for gross abnormalities.
Statistics:
Evaluation of Results and Statistical Analysis
The findings of this study were evaluated in terms of the observed effects, the necropsy and the microscopic findings.
Parameters like body weight gain and food consumption were calculated for each animal as the difference in weight measured from one week to the next. Mean body weights are also presented as figures. The relative organ weights were calculated in relation to the body weight (measured at necropsy) and are presented as percentage.
A statistical assessment of the results of the body weight. food consumption, parameters of haematology, blood coagulation and clinical biochemistry and absolute and relative organ weights were performed for each gender by comparing values of dosed with control animals of the main groups using a one-way ANOVA and a post-hoc Dunnett Test. These statistics were performed with GraphPad Prism 5.01 software (p<0.05 was considered as statistically significant).
Reproductive indices:
Copulation index, fertility index and delivery index was calculated for each group. The calculations were made using the formula as below,
Copulation Index (%) = (No. of rats copulated / No.of pairs) X 100
Fertility Index (%) = (No. of females pregant / No.of females copulated) X 100
Offspring viability indices:
Delivery Index (%) = (No. of dams with live newborns / No.of pregnant dams) X 100

Results and discussion

Results: P0 (first parental generation)

General toxicity (P0)

Clinical signs:
effects observed, non-treatment-related
Description (incidence and severity):
There were no clinical signs recorded in male and female animals of treated groups that could be directly related to treatment. However, there were few clinical signs namely moving the bedding, nasal discharge (dark or reddish), salivation and piloerection seen occasionally and transiently during the study period in MD or HD group animals. These findings were considered to be due to local effect but not the systemic effect of the test item. These findings were considered not likely to be adverse. In addition, there was alopecia (on hind limb, forelimb, thorax and abdominal region) of few isolated animals of MD or HD groups, which was assumed to be incidental in origin. During the weekly detailed clinical observation, no significant changes or differences between the groups were found.
Mortality:
no mortality observed
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
In both males and females, no treatment related changes were noted for body weight and body weight change during the study period. Statistically there were significant increase in body weight change in female HD group during 2nd week of premating period when compared to control. In addition, there was lower mean body weight gain noted between days 1-7 of premating when compared to control without attaining the statistical significance. But, this increase or decrease in weight gain did not correlate with food intake during the same period. Hence, the changes were not considered likely to be adverse. There was decrease in body weight gain noted in female MD and HD groups during lactation period when compared to control. This decrease had no statistical significance and was not likely to be adverse.
Food consumption and compound intake (if feeding study):
no effects observed
Water consumption and compound intake (if drinking water study):
no effects observed
Ophthalmological findings:
no effects observed
Description (incidence and severity):
There were no ophthalmoscopic findings in any of the animals of this study.
Haematological findings:
no effects observed
Description (incidence and severity):
For details see 7.5.1.
Clinical biochemistry findings:
no effects observed
Description (incidence and severity):
For details see 7.5.1.
Behaviour (functional findings):
no effects observed
Description (incidence and severity):
No relevant effects of treatment were observed in any of the parameters of the functional observation battery before and at the end of the treatment period. There were no biologically relevant differences in body temperature between the groups.
Organ weight findings including organ / body weight ratios:
no effects observed
Histopathological findings: non-neoplastic:
no effects observed
Description (incidence and severity):
For details see 7.5.1.
Other effects:
no effects observed

Reproductive function / performance (P0)

Reproductive function: oestrous cycle:
no effects observed
Reproductive function: sperm measures:
no effects observed
Description (incidence and severity):
There were no treatment related changes noted for epididymal sperm motility (Motile Count %, Static Count % and Rapid Count %) measured in all animals of treated and control groups. The statistical analysis of epididymal sperm motility indicated no statistical significant difference between the treated and control groups.
Reproductive performance:
no effects observed
Description (incidence and severity):
No treatment related changes were noted for number of corpora lutea, number of implantation sites, number of live pups born on PND 0 and percentage of pre and post implantation loss in treated groups when compared to control. The statistical evaluation of data revealed no significant differences between the values of treated and control groups.

Effect levels (P0)

Key result
Dose descriptor:
NOAEL
Effect level:
>= 1 000 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: no adverse effects seen at highest test dose

Target system / organ toxicity (P0)

Key result
Critical effects observed:
no

Results: F1 generation

General toxicity (F1)

Clinical signs:
no effects observed
Description (incidence and severity):
No treatment related changes were noted for survival of the pups from PND 0 to PND 4 in treated groups when compared to control.
Mortality / viability:
no mortality observed
Description (incidence and severity):
One pup each in Control, LD and MD groups found dead or missing between PND 0 and 4. The missing pup was assumed to be cannibalized by the dam, this observation was considered to be incidental. No treatment related changes were considered for viability index (%).
Body weight and weight changes:
no effects observed
Sexual maturation:
not examined
Organ weight findings including organ / body weight ratios:
no effects observed
Gross pathological findings:
no effects observed
Description (incidence and severity):
No treatment related gross external findings were observed in any of the treated groups. However, there were few isolated findings noted in pups namely dry skin, dark spot on/ or dark abdomen, dark snout in control group; dark spot on forelimb and thoracic back, dark snout in LD group; dry skin in MD and HD groups. These findings were considered to be incidental in origin.
Histopathological findings:
not examined

Effect levels (F1)

Key result
Dose descriptor:
NOAEL
Generation:
F1
Effect level:
>= 1 000 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: No effects seen at highest dose tested

Target system / organ toxicity (F1)

Key result
Critical effects observed:
no

Overall reproductive toxicity

Key result
Reproductive effects observed:
no

Applicant's summary and conclusion

Conclusions:
Repeated dose administration of Yttrium Oxide to male (minimum 28 days) and female (maximum 54 days) Wistar rats following OECD guideline 422 at dosages of 100, 300 and 1000 mg/kg body weight revealed neither mortalities nor findings of toxicological relevance in male and female animals. There were also no toxicologically relevant findings noted for reproductive and developmental parameters.

Based on these data, the no observed adverse effect level (NOAEL) for systemic and reproductive and developmental toxicity is considered to be 1000 mg/kg body weight/day. The NOEL for reproductive performance (mating and fertility) was considered to be 1000 mg/kg/day and the NOEL for toxic effects on progeny was 1000 mg/kg/day.
Executive summary:

A repeated dose study was performed according to OECD guideline 422 with Yttrium Oxide. Male (minimum 28 days) and female (maximum 54 days) Wistar rats were dosed at dosages of 100, 300 and 1000 mg/kg body weight/day. There were no changes considered to be related to treatment noted for organ weight in both males and females when compared to corresponding control. However, there was statistically significant increase in relative weight of left kidney weight in male treated (LD, MD and HD) groups, but not total kidney weight. This change in left kidney weight, in the absence of histological changes was not considered to have toxicological relevance.

No test item-related effects were noted on male and female reproductive organs in any of the treatment groups. The statistical analysis of epididymal sperm motility indicated no statistical significant difference between the treated and control groups. No treatment related changes were noted for the precoital interval and duration of gestation in treated groups when compared to control. All pregnancies resulted in normal births.

There were no treatment related changes noted for copulation index (%), fertility index (%), delivery index (%) and viability index (%) in treated groups when compared to corresponding control group.

Successful mating resulted 100% pregnancy rates in C and HD groups and 90% pregnancy rates in LD and MD groups. Histologically, they showed physiological sexual cycling, and their unsuccessful mating was considered unrelated to the treatment. The statistical evaluation of pre and post-natal data revealed no significant differences between the values of treated and control groups. No treatment related no significant changes were noted for survival of the pups and for viability index (%).

Based on these data, the no observed adverse effect level (NOAEL) for systemic and reproductive and developmental toxicity is considered to be 1000 mg/kg body weight/day. The NOEL for reproductive performance (mating and fertility) was considered to be 1000 mg/kg/day and the NOEL for toxic effects on progeny was 1000 mg/kg/day.