Cerium terbium magnesium undecaaluminum nonadecaoxide (hexagonal) ((Ce, Tb)MgAl11O19) and aluminium oxide (Al2O3) from calcination of Al2O3, CeO2, MgO and Tb4O7

Repeated dose toxicity: oral

Administrative data

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

Study period:

May 2015 - August 2015

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Cross-reference

Cross-reference 1

Reason / purpose for cross-reference:

reference to same study

Reference

Endpoint:

screening for reproductive / developmental toxicity

Type of information:

experimental study

Adequacy of study:

key study

Study period:

May 2015 - August 2015

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:

OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)

Version / remarks:

1996

Deviations:

no

Qualifier:

according to guideline

Guideline:

other: OPPTS 870.3650, 2000

Deviations:

no

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 421 (Reproduction / Developmental Toxicity Screening Test)

Version / remarks:

1995

Deviations:

no

Qualifier:

equivalent or similar to guideline

Guideline:

other: other guideline: OPPTS 870.3550, 2000

Deviations:

no

Qualifier:

equivalent or similar to guideline

Guideline:

other: EU Method B.7 (Repeated Dose (28 Days) Toxicity (Oral)), 2008

Deviations:

no

Principles of method if other than guideline:

also essentially conform the following guidelines:
-OECD Guidelines for Testing of Chemicals, Guideline 407, Repeated Dose 28-day Oral Toxicity Study in Rodents, October 2008.
-The United States EPA Health Effects Test Guidelines, OPPTS 870.3050, Repeated dose 28-day oral toxicity study in rodents, July 2000.

GLP compliance:

yes

Limit test:

no

Species:

rat

Strain:

other: Crl:WI(Han)

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Deutschland, Sulzfeld, Germany
- Age at study initiation: Approximately 11-12 weeks
- Weight at study initiation: males 297 - 355 grams, females 195 - 236 grams
- Fasting period before study: no
- Housing: Animals were housed in groups of 5 animals/sex/cage in Macrolon plastic cages during premating; For mating, females were caged together with males on a one-to-one-basis in; after mating males were housed in their home cage with a maximum of 5 animals/cage. Females were individually housed in Macrolon plastic cages; Pups were kept with the dam until termination in Macrolon plastic cages. Sterilized sawdust as bedding material and paper as cage-enrichment/nesting material were supplied. During locomotor activity monitoring, animals were housed individually in a Hitemp polycarbonate cage without cage-enrichment, bedding material, food and water.
- Diet: Free access to pelleted rodent diet (SM R/M-Z from SSNIFF® Spezialdiäten GmbH, Soest, Germany).
- Water: Free access to tap-water.
- Acclimation period: at least 5 days prior to start of treatment

ENVIRONMENTAL CONDITIONS
- Temperature: set to maintain 18 to 24°C
- Humidity: set to maintain 40 to 70%,
- Air changes (per hr): at least 10
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: 26 June 2015 To: 11 August 2015

Route of administration:

oral: gavage

Vehicle:

CMC (carboxymethyl cellulose)

Remarks:

1% in water

Details on exposure:

PREPARATION OF DOSING SOLUTIONS:
Formulations (w/v) were prepared daily within 6 hours prior to dosing and homogenized to a visually acceptable level. No correction was made for the purity/composition of the test substance.
Dose volume is 5 mL/kg bw. Actual dose volumes were calculated according to the latest body weight.

VEHICLE
- Justification for use and choice of vehicle (if other than water): Based on trial formulations performed at WIL Research Europe.

Details on mating procedure:

- Impregnation procedure: cohoused
- If cohoused:
- M/F ratio per cage: 1/1
- Length of cohabitation: A maximum of 14 days was allowed for mating.
- Verification of same strain and source of both sexes: yes
- Proof of pregnancy: vaginal plug / sperm in vaginal smear referred to as day 0 post-coitum.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Analyses were conducted on a single occasion during the treatment phase, according to a validated method (Project 507669). Samples of formulations were analyzed for homogeneity (highest and lowest concentration) and accuracy of preparation (all concentrations). The accuracy of preparation was considered acceptable if the mean measured concentrations were 85-115% of the target concentration. Homogeneity was demonstrated if the coefficient of variation was ≤ 10%.
Analysis of stability of the test substance under test conditions was not performed. Due to its extremely inert structure, the test substance will not disintegrate under the test conditions used in this study.

Duration of treatment / exposure:

Males were exposed for 31 days, i.e. 2 weeks prior to mating, during mating, and up to the day prior to scheduled necropsy. Females were exposed for 41-46 days, i.e. during 2 weeks prior to mating, during mating, during post-coitum, and during at least 4 days of lactation (up to the day prior to scheduled necropsy).

Frequency of treatment:

Once daily for 7 days per week

Remarks:

Doses / Concentrations:
100, 300, 1000, 1000 mg/kg bw/day
Basis:
actual ingested

No. of animals per sex per dose:

10

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: Dose levels were based on results of a 10-day dose range finding study (Project 507667)
An additional high dose group was included to test two different batches of the test substance.

Parental animals: Observations and examinations:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: At least twice daily.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Daily from treatment onwards up to the day prior to necropsy. Once prior to start of treatment and at weekly intervals during the treatment period this was also performed outside the home cage in a standard arena.

BODY WEIGHT: Yes
- Time schedule for examinations: Males and females were weighed on the first day of exposure and weekly thereafter. Mated females were weighed on Days 0, 4, 7, 11, 14, 17 and 20 post-coitum and during lactation on Days 1 and 4.

FOOD CONSUMPTION:
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes

WATER CONSUMPTION: No
Subjective appraisal was maintained during the study, but no quantitative investigation was introduced as no treatment related effect was suspected.

OTHER: HAEMATOLOGY, CLINICAL CHEMISTRY, FOB

Oestrous cyclicity (parental animals):

yes

Sperm parameters (parental animals):

Parameters examined in male parental generation: testis weight, epididymis weight, staging of spermatogenesis, sperm morphology

Litter observations:

STANDARDISATION OF LITTERS
- Performed on day 4 postpartum: no

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

GROSS EXAMINATION OF DEAD PUPS:
yes, for external abnormalities; The stomach of pups not surviving to the scheduled necropsy date was examined for the presence of milk, if possible.

Postmortem examinations (parental animals):

SACRIFICE
- Male animals: All surviving animals following completion of the mating period (a minimum of 28 days of dose administration).
- Maternal animals: All surviving females which delivered lactation Days 5-7; Females which failed to deliver post-coitum Days 25 or 27; Female with total litter loss within 24 hours of litter loss.

GROSS PATHOLOGY: Yes, according to guidelines

HISTOPATHOLOGY: Yes, according to guidelines
organ weights: (5/sex/group) adrenal glands, spleen, brain, testes, epididymides, thymus, heart, uterus (including cervix), kidneys, prostate, liver, seminal vesicles including coagulating glands, ovaries, thyroid including parathyroid; all remaining males: epididymides, testes.

Postmortem examinations (offspring):

SACRIFICE
- The F1 offspring was sacrificed at 5-7 days of age.

GROSS NECROPSY
- Gross necropsy

Statistics:

The following statistical methods were used to analyse the data:
- If the variables could be assumed to follow a normal distribution, the Dunnett-test (many-to-one t-test) based on a pooled variance estimate was applied for the comparison of the treated groups and the control groups for each sex.
- The Steel-test (many-to-one rank test) was applied if the data could not be assumed to follow a normal distribution.
- The Fisher Exact-test was applied to frequency data.
- The Kruskal-Wallis nonparametric ANOVA test was applied to motor activity data to determine intergroup differences.
All tests were two-sided and in all cases p < 0.05 was accepted as the lowest level of significance.
Group means were calculated for continuous data and medians were calculated for discrete data (scores) in the summary tables. Test statistics were calculated on the basis of exact values for means and pooled variances.

Reproductive indices:

For each group, the following calculations were performed:

Mating index (%) (Number of females mated/ Number of females paired)x 100

Fertility index (%) N(umber of pregnant females/ Number of females paired) x 100

Conception index (%) (Number of pregnant females/ Number of females mated) x 100

Gestation index (%) (Number of females bearing live pups/ Number of pregnant females) x 100

Duration of gestation Number of days between confirmation of mating and the beginning of parturition

Offspring viability indices:

For each group, the following calculations were performed:

Percentage live males at First Litter Check (Number of live male pups at First Litter Check/ Number of live pups at First Litter Check) x 100

Percentage live females at First Litter Check (Number of live female pups at First Litter Check/ Number of live pups at First Litter Check) x 100

Percentage of postnatal loss (Number of dead pups before planned necropsy/ Number of live pups at First Litter Check) x 100

Viability index (Number of live pups before planned necropsy/ Number of pups born alive) x 100

Clinical signs:

no effects observed

Body weight and weight changes:

no effects observed

Food consumption and compound intake (if feeding study):

no effects observed

Organ weight findings including organ / body weight ratios:

no effects observed

Histopathological findings: non-neoplastic:

no effects observed

Reproductive function: oestrous cycle:

no effects observed

Reproductive function: sperm measures:

no effects observed

Reproductive performance:

no effects observed

CLINICAL SIGNS AND MORTALITY (PARENTAL ANIMALS):
There were two premature decedents in this study. None of these deaths was considered to be test substance related (one female dosed 300 mg/kg bw/day with total litter loss, one high dose group female (group 4) was found dead on lactation Day 1). There were no clinical signs of toxicity during the observation period. One Group 1 male was noted with hunched posture, piloerection and a lean appearance towards the end of the treatment period. Since this was a male of the control group, these findings were by no means test substance related.

BODY WEIGHT AND FOOD CONSUMPTION (PARENTAL ANIMALS):
Body weights and body weight gain of treated animals remained in the same range as controls over the treatment period. No toxicologically relevant changes in food consumption before or after allowance for body weight were noted.

REPRODUCTIVE FUNCTION: ESTROUS CYCLE (PARENTAL ANIMALS):
no adverse effects observed

REPRODUCTIVE FUNCTION: SPERM MEASURES (PARENTAL ANIMALS):
Spermatogenic staging profiles were normal for all males examined.

REPRODUCTIVE PERFORMANCE (PARENTAL ANIMALS):
There were no treatment-related findings on the reproduction parameters observed. The mating index, fertility and conception indices, precoital time, and number of corpora lutea and implantation sites were unaffected by treatment.
There was one control couple, two couples treated at 100 mg/kg bw/day, one couple treated at 300 mg/kg bw/day and three couples at 1000 mg/kg bw/day without offspring. There was one couple treated at 300 mg/kg bw/day with total litter loss. No abnormalities were seen in the reproductive organs (and mammary gland), which could account for their lack of offspring. There were no morphological findings in the reproductive organs of either sex which could be attributed to the test item and spermatogenic staging profiles were normal for all males examined.

ORGAN WEIGHTS (PARENTAL ANIMALS):
There were no test item-related alterations in organ weights.
The lower seminal vesicle organ weight (absolute and relative to body weight) noted for males at 100 mg/kg bw/day as compared to the concurrent control group was considered not to be a sign of toxicity. No dose-related trend could be established and no corroborative findings were present either macroscopically or microscopically.

GROSS PATHOLOGY (PARENTAL ANIMALS):
There were no test item-related gross observations.
All recorded macroscopic findings were within the range of background observations encountered in rats of this age and strain.

HISTOPATHOLOGY (PARENTAL ANIMALS):
There were no test item-related microscopic observations.
All of the recorded microscopic findings were within the range of background pathology encountered in rats of this age and strain. There was no test item-related alteration in the prevalence, severity, or histologic character of those incidental tissue alterations.

Key result

Dose descriptor:

NOAEL

Remarks:

reproduction

Effect level:

>= 1 000 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: no test substance related adverse effects observed up to and including 1000 mg/kg bw/day, the highest dose tested

Key result

Dose descriptor:

NOAEL

Remarks:

parental

Effect level:

>= 1 000 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: no test substance related adverse effects observed up to and including 1000 mg/kg bw/day, the highest dose tested

Clinical signs:

no effects observed

Mortality / viability:

no mortality observed

Body weight and weight changes:

no effects observed

Sexual maturation:

not examined

Organ weight findings including organ / body weight ratios:

not examined

Gross pathological findings:

no effects observed

Histopathological findings:

not examined

VIABILITY (OFFSPRING):
At first litter check, in total three pups from two different litters at 300 mg/kg bw/day were found dead. In the absence of any dead pups at the higher dose level of 1000 mg/kg bw/day, no toxicological significance was attached to this finding. During lactation, one pup of the control group and two pups at 300 mg/kg bw/day went missing. These missing pups were most likely cannibalised. No toxicological relevance was attributed to these missing pups since the mortality incidence did not show a dose related trend and remained within the range considered normal for pups of this age. All three pups of one 1000 mg/kg bw/day litter had to be euthanized on lactation Day 1 due to the spontaneous death of their mother.

CLINICAL SIGNS (OFFSPRING):
One pup at 300 mg/kg bw/day that went missing on lactation Day 2 had no milk in its stomach at fi rst litter check. Incidental clinical symptoms of pups surviving until scheduled necropsy consisted of blue spot on the nose and scabbing of the cheek. The nature and incidence of these clinical signs remained within the range considered normal for pups of this age, and were therefore considered to be of no toxicological relevance.

BODY WEIGHT (OFFSPRING):
Body weights of pups were considered to have been unaffected by treatment

GROSS PATHOLOGY (OFFSPRING):
Incidental macroscopic findings of pups that were found dead included (beginning) autolysis. Incidental macroscopic findings among surviving pups included scabbing of the cheek. The nature and incidence of these findings remained within the range considered normal for pups of this age, and were therefore considered to be of no toxicological relevance.

Key result

Dose descriptor:

NOAEL

Generation:

F1

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 observed up to and including 1000 mg/kg bw/day, the highest dose tested

Reproductive effects observed:

not specified

Conclusions:

A reliable OECD 422 study is available, in which no test substance related adverse effects were observed up to and including 1000 mg/kg bw/day, the highest dose tested.

Executive summary:

In an OECD 422 study, CAT was administered by daily oral gavage to male and female Wistar Han rats at dose levels of 100, 300 and 1000 mg/kg bw/day (batch B) and 1000 mg/kg bw/day (batch C). Males were exposed for 2 weeks prior to mating, during mating, and up to termination (for 31 days). The females were exposed for 2 weeks prior to mating, during mating, during post-coitum, and at least 4 days of lactation (for 41-46 days). Formulation analysis showed that the formulations were prepared accurately and homogenously.

No parental toxicity was observed up to and including the highest dose level tested (1000 mg/kg bw/day of batch B or C) as evidenced by the absence of clinical signs of toxicity or adverse changes in functional observational results, body weight (gain), food consumption, haematology and clinical biochemistry parameters, organ weights, macroscopic findings, and microscopic findings. No reproductive toxicity was observed up to and including the highest dose level tested (1000 mg/kg bw/day for both batches). No treatment-related changes were noted in any of the reproductive parameters investigated in this study (i.e. mating, fertility and conception indices, precoital time, and numbers of corpora lutea and implantation sites, spermatogenic profiling, and histopathological examination of reproductive organs). The observed number of pregnant females (n=7) in the 1000 mg/kg bw/day dose group (batch C) was just below the minimum of 8 advised by the guideline. This finding was considered a chance finding and not treatment related, as no abnormalities were seen in the reproductive organs of either sex, which could account for their lack of offspring. In addition, spermatogenic staging profiles were normal for all males examined. All 7 pregnant females delivered healthy pups and litter sizes were normal. Sufficient offspring was produced to assure a meaningful evaluation of possible developmental (from implantation onwards) toxicity of the test substance.

In conclusion, treatment with CAT by oral gavage in male and female Wistar Han rats at dose levels of 100, 300 and 1000 mg/kg bw/day (batch B) and 1000 mg/kg bw/day (batch C) did not result in parental reproductive or developmental adverse effects. Based on these results, a parental, reproductive and developmental No Observed Adverse Effect Level (NOAEL) of at least 1000 mg/kg bw/day was derived for both batches (B and C).

Cross-reference 2

Reason / purpose for cross-reference:

reference to same study

Reference

Endpoint:

developmental toxicity

Type of information:

experimental study

Adequacy of study:

key study

Study period:

May 2015 - August 2015

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: OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test), 1996

Deviations:

no

Qualifier:

according to guideline

Guideline:

other: OPPTS 870.3650, 2000

Deviations:

no

Qualifier:

equivalent or similar to guideline

Guideline:

other: OECD 421, 1995

Deviations:

no

Qualifier:

equivalent or similar to guideline

Guideline:

other: OPPTS 870.3550, 2000

Deviations:

no

Qualifier:

equivalent or similar to guideline

Guideline:

other: EU Method B.7 (Repeated Dose (28 Days) Toxicity (Oral)), 2008

Deviations:

no

Principles of method if other than guideline:

also essentially conform the following guidelines:
-OECD Guidelines for Testing of Chemicals, Guideline 407, Repeated Dose 28-day Oral Toxicity Study in Rodents, October 2008.
-The United States EPA Health Effects Test Guidelines, OPPTS 870.3050, Repeated dose 28-day oral toxicity study in rodents, July 2000.

GLP compliance:

yes

Limit test:

no

Species:

rat

Strain:

other: Crl:WI(Han)

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Deutschland, Sulzfeld, Germany
- Age at study initiation: Approximately 11-12 weeks
- Weight at study initiation: males 297 - 355 grams, females 195 - 236 grams
- Fasting period before study: no
- Housing: Animals were housed in groups of 5 animals/sex/cage in Macrolon plastic cages during premating; For mating, females were caged together with males on a one-to-one-basis in; after mating males were housed in their home cage with a maximum of 5 animals/cage. Females were individually housed in Macrolon plastic cages; Pups were kept with the dam until termination in Macrolon plastic cages. Sterilized sawdust as bedding material and paper as cage-enrichment/nesting material were supplied. During locomotor activity monitoring, animals were housed individually in a Hitemp polycarbonate cage without cage-enrichment, bedding material, food and water.
- Diet: Free access to pelleted rodent diet (SM R/M-Z from SSNIFF® Spezialdiäten GmbH, Soest, Germany).
- Water: Free access to tap-water.
- Acclimation period: at least 5 days prior to start of treatment

ENVIRONMENTAL CONDITIONS
- Temperature: set to maintain 18 to 24°C
- Humidity: set to maintain 40 to 70%,
- Air changes (per hr): at least 10
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: 26 June 2015 To: 11 August 2015

Route of administration:

oral: gavage

Vehicle:

CMC (carboxymethyl cellulose)

Remarks:

1% in water

Details on exposure:

PREPARATION OF DOSING SOLUTIONS:
Formulations (w/v) were prepared daily within 6 hours prior to dosing and homogenized to a visually acceptable level. No correction was made for the purity/composition of the test substance.
Dose volume is 5 mL/kg bw. Actual dose volumes were calculated according to the latest body weight.

VEHICLE
- Justification for use and choice of vehicle (if other than water): Based on trial formulations performed at WIL Research Europe.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Analyses were conducted on a single occasion during the treatment phase, according to a validated method (Project 507669). Samples of formulations were analyzed for homogeneity (highest and lowest concentration) and accuracy of preparation (all concentrations). The accuracy of preparation was considered acceptable if the mean measured concentrations were 85-115% of the target concentration. Homogeneity was demonstrated if the coefficient of variation was ≤ 10%.
Analysis of stability of the test substance under test conditions was not performed. Due to its extremely inert structure, the test substance will not disintegrate under the test conditions used in this study.

Details on mating procedure:

- Impregnation procedure: cohoused
- If cohoused:
- M/F ratio per cage: 1/1
- Length of cohabitation: A maximum of 14 days was allowed for mating.
- Verification of same strain and source of both sexes: yes
- Proof of pregnancy: vaginal plug / sperm in vaginal smear referred to as day 0 post-coitum.

Duration of treatment / exposure:

Males were exposed for 31 days, i.e. 2 weeks prior to mating, during mating, and up to the day prior to scheduled necropsy. Females were exposed for 41-46 days, i.e. during 2 weeks prior to mating, during mating, during post-coitum, and during at least 4 days of lactation (up to the day prior to scheduled necropsy).
Pups were not dosed directly but could have potentially exposed to the test substance in utero, via maternal milk or from exposure to maternal urine/faeces.

Frequency of treatment:

Once daily for 7 days per week

No. of animals per sex per dose:

10

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: Dose levels were based on results of a 10-day dose range finding study (Project 507667)
An additional high dose group was included to test two different batches of the test substance.

Maternal examinations:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: At least twice daily.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Daily from treatment onwards up to the day prior to necropsy. Once prior to start of treatment and at weekly intervals during the treatment period this was also performed outside the home cage in a standard arena.

BODY WEIGHT: Yes
- Time schedule for examinations: Males and females were weighed on the first day of exposure and weekly thereafter. Mated females were weighed on Days 0, 4, 7, 11, 14, 17 and 20 post-coitum and during lactation on Days 1 and 4.

FOOD CONSUMPTION:
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes

WATER CONSUMPTION: No
Subjective appraisal was maintained during the study, but no quantitative investigation was introduced as no treatment related effect was suspected.

POST-MORTEM EXAMINATIONS: YES
GROSS PATHOLOGY: Yes, according to guidelines
HISTOPATHOLOGY: Yes, according to guidelines
organ weights: (5/sex/group) adrenal glands, spleen, brain, testes, epididymides, thymus, heart, uterus (including cervix), kidneys, prostate, liver, seminal vesicles including coagulating glands, ovaries, thyroid including parathyroid; all remaining males: epididymides, testes.

OTHER: HAEMATOLOGY, CLINICAL CHEMISTRY, FOB

Ovaries and uterine content:

The ovaries and uterine content was examined after termination: Yes / No / No data
Examinations included:
- Gravid uterus weight: No
- Number of corpora lutea: Yes
- Number of implantations: Yes
- Number of early resorptions: No
- Number of late resorptions: No

Fetal examinations:

- External examinations: Yes: all per litter
- Soft tissue examinations: No
- Skeletal examinations: No
- Head examinations: No

Statistics:

The following statistical methods were used to analyse the data:
- If the variables could be assumed to follow a normal distribution, the Dunnett-test (many-to-one t-test) based on a pooled variance estimate was applied for the comparison of the treated groups and the control groups for each sex.
- The Steel-test (many-to-one rank test) was applied if the data could not be assumed to follow a normal distribution.
- The Fisher Exact-test was applied to frequency data.
- The Kruskal-Wallis nonparametric ANOVA test was applied to motor activity data to determine intergroup differences.
All tests were two-sided and in all cases p < 0.05 was accepted as the lowest level of significance.
Group means were calculated for continuous data and medians were calculated for discrete data (scores) in the summary tables. Test statistics were calculated on the basis of exact values for means and pooled variances.

Indices:

For each group, the following calculations were performed:

Percentage live males at First Litter Check (Number of live male pups at First Litter Check/ Number of live pups at First Litter Check) x 100

Percentage live females at First Litter Check (Number of live female pups at First Litter Check/ Number of live pups at First Litter Check) x 100

Percentage of postnatal loss (Number of dead pups before planned necropsy/ Number of live pups at First Litter Check) x 100

Viability index (Number of live pups before planned necropsy/ Number of pups born alive) x 100

Details on maternal toxic effects:

Maternal toxic effects:no effects

Details on maternal toxic effects:
No parental toxicity was observed up to the highest dose level tested (1000 mg/kg bw/day of batch B or C) as evidenced by the absence of clinical signs of toxicity or adverse changes in functional observational results, body weight (gain), food consumption, haematology and clinical biochemistry parameters, organ weights, macroscopic findings, and microscopic findings. No treatment-related changes were noted in any of the reproductive parameters investigated in this study (i.e. mating, fertility and conception indices, precoital time, and numbers of corpora lutea and implantation sites, and histopathological examination of reproductive organs).

Key result

Dose descriptor:

NOAEL

Effect level:

>= 1 000 mg/kg bw/day (actual dose received)

Based on:

test mat.

Basis for effect level:

other: no substance related adverse effects observed up to and including 1000 mg/kg bw/day, the highest dose tested

Details on embryotoxic / teratogenic effects:

Embryotoxic / teratogenic effects:no effects

Details on embryotoxic / teratogenic effects:
No toxicologically relevant effects on gestation index and duration, parturition, maternal care and early postnatal pup development (mortality, clinical signs, body weight and macroscopy) were observed. The numbers of females with living pups on Day 1 of lactation were 9, 8, 8, 10 and 7 at 0, 100, 300 and 1000 mg/kg bw/day (batch B) and 1000 mg/kg bw/day (batch C), respectively. The gestation index and duration of gestation were unaffected by treatment. No signs of difficult or prolonged parturition were noted among the pregnant females. Examination of cage debris of pregnant females revealed no signs of abortion or premature birth. No deficiencies in maternal care were observed. Number of dead and living pups at first litter check, postnatal loss, viability index and sex ratio were unaffected by treatment, and clinical signs, body weight and external macroscopy did not reveal treatment-related findings.

VIABILITY (OFFSPRING):
At first litter check, in total three pups from two different litters at 300 mg/kg bw/day were found dead. In the absence of any dead pups at the higher dose level of 1000 mg/kg bw/day, no toxicological significance was attached to this finding. During lactation, one pup of the control group and two pups at 300 mg/kg bw/day went missing. These missing pups were most likely cannibalised. No toxicological relevance was attributed to these missing pups since the mortality incidence did not show a dose related trend and remained within the range considered normal for pups of this age. All three pups of one 1000 mg/kg bw/day litter had to be euthanized on lactation Day 1 due to the spontaneous death of their mother.

CLINICAL SIGNS (OFFSPRING):
One pup at 300 mg/kg bw/day that went missing on lactation Day 2 had no milk in its stomach at fi rst litter check. Incidental clinical symptoms of pups surviving until scheduled necropsy consisted of blue spot on the nose and scabbing of the cheek. The nature and incidence of these clinical signs remained
within the range considered normal for pups of this age, and were therefore considered to be of no toxicological relevance.

BODY WEIGHT (OFFSPRING):
Body weights of pups were considered to have been unaffected by treatment

GROSS PATHOLOGY (OFFSPRING):
Incidental macroscopic findings of pups that were found dead included (beginning) autolysis. Incidental macroscopic findings among surviving pups included scabbing of the cheek. The nature and incidence of these findings remained within the range considered normal for pups of this age, and were therefore considered to be of no toxicological relevance.

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 observed up to and including 1000 mg/kg bw/day, the highest dose tested

Developmental effects observed:

no

Conclusions:

A reliable OECD 422 study is available, in which no test substance related adverse effects were observed up to and including 1000 mg/kg bw/day, the highest dose tested.

Executive summary:

In an OECD 422 study, CAT was administered by daily oral gavage to male and female Wistar Han rats at dose levels of 100, 300 and 1000 mg/kg bw/day (batch B) and 1000 mg/kg bw/day (batch C). Males were exposed for 2 weeks prior to mating, during mating, and up to termination (for 31 days). The females were exposed for 2 weeks prior to mating, during mating, during post-coitum, and at least 4 days of lactation (for 41-46 days). Formulation analysis showed that the formulations were prepared accurately and homogenously.

No parental toxicity was observed up to and including the highest dose level tested (1000 mg/kg bw/day of batch B or C) as evidenced by the absence of clinical signs of toxicity or adverse changes in functional observational results, body weight (gain), food consumption, haematology and clinical biochemistry parameters, organ weights, macroscopic findings, and microscopic findings. No reproductive toxicity was observed up to and including the highest dose level tested (1000 mg/kg bw/day for both batches). No treatment-related changes were noted in any of the reproductive parameters investigated in this study (i.e. mating, fertility and conception indices, precoital time, and numbers of corpora lutea and implantation sites, spermatogenic profiling, and histopathological examination of reproductive organs). No toxicologically relevant effects on gestation index and duration, parturition, maternal care and early postnatal pup development (mortality, clinical signs, body weight and macroscopy) were observed.

In conclusion, treatment with CAT by oral gavage in male and female Wistar Han rats at dose levels of 100, 300 and 1000 mg/kg bw/day (batch B) and 1000 mg/kg bw/day (batch C) did not result in parental, reproductive or developmental adverse effects. Based on these results, a parental, reproductive and developmental No Observed Adverse Effect Level (NOAEL) of at least 1000 mg/kg bw/day was derived for both batches (B and C).

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2015

Report date:

2015

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)

Version / remarks:

1996

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

other: OPPTS 870.3650, 2000

Deviations:

no

Test guideline 3

Qualifier:

equivalent or similar to guideline

Guideline:

other: OECD 421, 1995

Deviations:

no

Test guideline 4

Qualifier:

equivalent or similar to guideline

Guideline:

other: OPPTS 870.3550, 2000

Deviations:

no

Test guideline 5

Qualifier:

equivalent or similar to guideline

Guideline:

EU Method B.7 (Repeated Dose (28 Days) Toxicity (Oral))

Version / remarks:

2008

Deviations:

no

Principles of method if other than guideline:

also essentially conform the following guidelines:
-OECD Guidelines for Testing of Chemicals, Guideline 407, Repeated Dose 28-day Oral Toxicity Study in Rodents, October 2008.
-The United States EPA Health Effects Test Guidelines, OPPTS 870.3050, Repeated dose 28-day oral toxicity study in rodents, July 2000.

GLP compliance:

yes

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Cerium terbium magnesium undecaaluminum nonadecaoxide

Molecular formula:

Ce0.5Tb0.5MgAl11O19 -Ce0.7Tb0.3MgAl11O19

IUPAC Name:

Cerium terbium magnesium undecaaluminum nonadecaoxide

Constituent 2

Reference substance name:

Aluminium oxide

EC Number:

215-691-6

EC Name:

Aluminium oxide

Cas Number:

1344-28-1

Molecular formula:

Al2O3

IUPAC Name:

aluminium trioxide

Test material form:

solid: particulate/powder

Details on test material:

- Name of test material (as cited in study report): CAT
- Substance type: white powder
- Storage condition of test material: at room temperature

Test animals

Species:

rat

Strain:

other: Crl:WI(Han)

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Deutschland, Sulzfeld, Germany
- Age at study initiation: Approximately 11-12 weeks
- Weight at study initiation: males 297 - 355 grams, females 195 - 236 grams
- Fasting period before study: no
- Housing: Animals were housed in groups of 5 animals/sex/cage in Macrolon plastic cages during premating; For mating, females were caged together with males on a one-to-one-basis in; after mating males were housed in their home cage with a maximum of 5 animals/cage. Females were individually housed in Macrolon plastic cages; Pups were kept with the dam until termination in Macrolon plastic cages. Sterilized sawdust as bedding material and paper as cage-enrichment/nesting material were supplied. During locomotor activity monitoring, animals were housed individually in a Hitemp polycarbonate cage without cage-enrichment, bedding material, food and water.
- Diet: Free access to pelleted rodent diet (SM R/M-Z from SSNIFF® Spezialdiäten GmbH, Soest, Germany).
- Water: Free access to tap-water.
- Acclimation period: at least 5 days prior to start of treatment

ENVIRONMENTAL CONDITIONS
- Temperature: set to maintain 18 to 24°C
- Humidity: set to maintain 40 to 70%,
- Air changes (per hr): at least 10
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: 26 June 2015 To: 11 August 2015

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

CMC (carboxymethyl cellulose)

Remarks:

1% in water

Details on oral exposure:

PREPARATION OF DOSING SOLUTIONS:
Formulations (w/v) were prepared daily within 6 hours prior to dosing and homogenized to a visually acceptable level. No correction was made for the purity/composition of the test substance.
Dose volume is 5 mL/kg bw. Actual dose volumes were calculated according to the latest body weight.

VEHICLE
- Justification for use and choice of vehicle (if other than water): Based on trial formulations performed at WIL Research Europe.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Analyses were conducted on a single occasion during the treatment phase, according to a validated method (Project 507669). Samples of formulations were analyzed for homogeneity (highest and lowest concentration) and accuracy of preparation (all concentrations). The accuracy of preparation was considered acceptable if the mean measured concentrations were 85-115% of the target concentration. Homogeneity was demonstrated if the coefficient of variation was ≤ 10%.
Analysis of stability of the test substance under test conditions was not performed. Due to its extremely inert structure, the test substance will not disintegrate under the test conditions used in this study.

Duration of treatment / exposure:

Males were exposed for 31 days, i.e. 2 weeks prior to mating, during mating, and up to the day prior to scheduled necropsy. Females were exposed for 41-46 days, i.e. during 2 weeks prior to mating, during mating, during post-coitum, and during at least 4 days of lactation (up to the day prior to scheduled necropsy).

Frequency of treatment:

Once daily for 7 days per week.

Doses / concentrations

Remarks:

Doses / Concentrations:
100, 300, 1000, 1000 mg/kg bw /day
Basis:
actual ingested

No. of animals per sex per dose:

10

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: Dose levels were based on results of a 10-day dose range finding study (Project 507667)
An additional high dose group was included to test two different batches of the test substance.

Examinations

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: At least twice daily.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Daily from treatment onwards up to the day prior to necropsy. Once prior to start of treatment and at weekly intervals during the treatment period this was also performed outside the home cage in a standard arena.

BODY WEIGHT: Yes
- Time schedule for examinations: Males and females were weighed on the first day of exposure and weekly thereafter. Mated females were weighed on Days 0, 4, 7, 11, 14, 17 and 20 post-coitum and during lactation on Days 1 and 4.

FOOD CONSUMPTION:
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes

FOOD EFFICIENCY:
- Body weight gain in kg/food consumption in kg per unit time X 100 calculated as time-weighted averages from the consumption and body weight gain data: No data

WATER CONSUMPTION: No
Subjective appraisal was maintained during the study, but no quantitative investigation was introduced as no treatment related effect was suspected.

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes
- Time schedule for collection of blood: at the end of the treatment period on the day of scheduled necropsy.
- Anaesthetic used for blood collection: isoflurane
- Animals fasted: Yes, o/n (maximum of 24 hours)
- How many animals: 5/sex/group
- Parameters according to guidelines were examined.

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: at the end of the treatment period on the day of scheduled necropsy.
- Animals fasted: Yes, o/n (maximum of 24 hours)
- How many animals: 5/sex/group
- Parameters according to guidelines were examined.

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: Yes (FOB)
- Time schedule for examinations: The selected males were tested during Week 4 of treatment and the selected females were tested towards the end of the scheduled lactation period (all before blood sampling). These tests were performed after observation for clinical signs.
- Dose groups that were examined: 5/sex/group
- Battery of functions tested: hearing ability, pupillary reflex and static righting reflex, fore- and hind-limb grip strength, locomotor activity (total movements and ambulations)

Sacrifice and pathology:

GROSS PATHOLOGY: Yes, according to guidelines
HISTOPATHOLOGY: Yes, according to guidelines
organ weights: (5/sex/group) adrenal glands, spleen, brain, testes, epididymides, thymus, heart, uterus (including cervix), kidneys, prostate, liver, seminal vesicles including coagulating glands, ovaries, thyroid including parathyroid; all remaining males: epididymides, testes.

Statistics:

The following statistical methods were used to analyse the data:
- If the variables could be assumed to follow a normal distribution, the Dunnett-test (many-to-one t-test) based on a pooled variance estimate was applied for the comparison of the treated groups and the control groups for each sex.
- The Steel-test (many-to-one rank test) was applied if the data could not be assumed to follow a normal distribution.
- The Fisher Exact-test was applied to frequency data.
- The Kruskal-Wallis nonparametric ANOVA test was applied to motor activity data to determine intergroup differences.
All tests were two-sided and in all cases p < 0.05 was accepted as the lowest level of significance.
Group means were calculated for continuous data and medians were calculated for discrete data (scores) in the summary tables. Test statistics were calculated on the basis of exact values for means and pooled variances.

Results and discussion

Results of examinations

Clinical signs:

no effects observed

Mortality:

no mortality observed

Body weight and weight changes:

no effects observed

Food efficiency:

no effects observed

Water consumption and compound intake (if drinking water study):

no effects observed

Ophthalmological findings:

not examined

Haematological findings:

no effects observed

Clinical biochemistry findings:

no effects observed

Urinalysis findings:

not examined

Behaviour (functional findings):

no effects observed

Organ weight findings including organ / body weight ratios:

no effects observed

Gross pathological findings:

no effects observed

Histopathological findings: non-neoplastic:

no effects observed

Histopathological findings: neoplastic:

no effects observed

Details on results:

CLINICAL SIGNS AND MORTALITY:
There were two premature decedents in this study. None of these deaths was considered to be test substance related (one female dosed 300 mg/kg bw/day with total litter loss, one high dose group female (group 4) was found dead on lactation Day 1). There were no clinical signs of toxicity during the observation period. One control male was noted with hunched posture, piloerection and a lean appearance towards the end of the treatment period. Since this was a male of the control group, these findings were by no means test substance related.

BODY WEIGHT AND WEIGHT GAIN:
Body weights and body weight gain of treated animals remained in the same range as controls over the treatment period.

FOOD CONSUMPTION:
No toxicologically relevant changes in food consumption before or after allowance for body weight were noted.

HAEMATOLOGY:
Haematological parameters of treated rats were considered not to have been affected by treatment. Slightly higher values of red blood cells, haemoglobin and haematocrit were noted for males at 100 mg/kg bw/day as compared to controls. These findings were considered to be of no toxicological relevance as they occurred in the absence of a treatment-related distribution and remained within the range considered normal for rats of this age and strain.

CLINICAL CHEMISTRY:
Clinical biochemistry parameters of treated rats were considered not to have been affected by treatment. Higher values of urea at 100 mg/kg bw/day (males), and glucose (males) and alanine aminotransferase (ALAT; females) at 300 mg/kg bw/day were considered to be of no toxicological significance as they occurred in the absence of a treatment-related distribution and remained within the range considered normal for rats of this age and strain.

NEUROBEHAVIOUR:
No toxicologically relevant effects on hearing ability, pupillary reflex, static righting reflex and grip strength were observed. One male (no. 15) at 100 mg/kg bw/day had no pupil reflex of his left eye. In the absence of corroborative findings at the higher doses, no toxicological relevance was attributed to this isolated finding. At the individual level, two males and one female at 300 mg/kg bw/day were noted with relatively high total numbers of total movements. No increased activity was seen for males and females treated at the higher dose of 1000 mg/kg bw/day. Therefore, this finding was not considered to be toxicologically relevant.

ORGAN WEIGHTS:
There were no test item-related alterations in organ weights.
The lower seminal vesicle organ weight (absolute and relative to body weight) noted for males at 100 mg/kg bw/day as compared to the concurrent control group was considered not to be a sign of toxicity. No dose-related trend could be established and no corroborative findings were present either macroscopically or microscopically.

GROSS PATHOLOGY:
There were no test item-related gross observations.
All recorded macroscopic findings were within the range of background observations encountered in rats of this age and strain.

HISTOPATHOLOGY: NON-NEOPLASTIC
There were no test item-related microscopic observations.
All of the recorded microscopic findings were within the range of background pathology encountered in rats of this age and strain. There was no test item-related alteration in the prevalence, severity, or histologic character of those incidental tissue alterations.

Effect levels

Key result

Dose descriptor:

NOAEL

Remarks:

parental

Effect level:

>= 1 000 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: no test substance related adverse effects observed up to and including 1000 mg/kg bw/day, the highest dose tested

Target system / organ toxicity

Critical effects observed:

not specified

Any other information on results incl. tables

Accuracy of preparation

In the Group 1 formulations, no test substance was detected. Mean recoveries/day were 83 -89%. One of the mean recoveries of the procedural recovery samples (83%) did not fall within the criterion of 85 − 115%. The rest of the mean recoveries of the procedural recovery samples fell within the criterion, however the mean recoveries were very close to the lower limit of the criterion. Therefore, the concentration of the test samples were corrected for the mean recovery of the procedural recovery samples which were pretreated on the same day.The corrected concentrations analysed in the formulations of Group 2, Group 3, Group 4 and Group 5 were in agreement with target concentrations (i.e. mean accuracies between 85% and 115%). Homogeneity The formulations of Group 2, Group 4 and Group 5 were homogeneous: 97%-106% (coefficient of variation ≤ 10%).

Applicant's summary and conclusion

Conclusions:

In the absence of adverse effects in parental rats in a repeated dose toxicity study with reproduction/developmental screening (OECD 422), the NOAEL is at least 1000 mg/kg bw/day for parental toxicity.

Executive summary:

In an OECD 422 study, CAT was administered by daily oral gavage to male and female Wistar Han rats at dose levels of 100, 300 and 1000 mg/kg bw/day (batch B) and 1000 mg/kg bw/day (batch C). Males were exposed for 2 weeks prior to mating, during mating, and up to termination (for 31 days). The females were exposed for 2 weeks prior to mating, during mating, during post-coitum, and at least 4 days of lactation (for 41-46 days). Formulation analysis showed that the formulations were prepared accurately and homogenously.

No parental toxicity was observed up to and including the highest dose level tested (1000 mg/kg bw/day of batch B or C) as evidenced by the absence of clinical signs of toxicity or adverse changes in functional observational results, body weight (gain), food consumption, haematology and clinical biochemistry parameters, organ weights, macroscopic findings, and microscopic findings. In conclusion, treatment with CAT by oral gavage in male and female Wistar Han rats at dose levels of 100, 300 and 1000 mg/kg bw/day(batch B) and 1000 mg/kg bw/day (batch C) did not result in parental adverse effects. Based on these results, a parental No Observed Adverse Effect Level (NOAEL) of at least 1000 mg/kg bw/day was derived for both batches (B and C).