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Description of key information

Repeated dose toxicity - oral

The key study (Papineau, 2018) was performed according to OECD guideline 422 and conform GLP requirements. In this study, ytterbium oxide was administered by oral gavage to male and female Sprague-Dawley rats, starting 2 weeks before mating, during mating and (for females) throughout gestation and until day 13 post-partum, at the dose levels of 110, 330 or 1000 mg/kg bw/day. In this study, the NOAEL for parental systemic toxicity was considered to be higher than or equal to 1000 mg/kg bw/day based on the absence of adverse findings related to the test item at the highest dose level. Based on these results, the test substance is not classified as STOT RE.

Repeated dose toxicity - inhalation/dermal

No key studies were identified for repeated dose toxicity after inhalation or dermal exposure.

A key study is available for the oral route of exposure. According to the REACH Regulation, only one route of exposure should be tested for repeated dose toxicity (Column 2, Annex VIII, Section 8.6.1). Therefore, it is not necessary to perform a repeated dose toxicity study via the inhalation or dermal route of exposure.

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records
Reference
Endpoint:
short-term repeated dose toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
6 July 2017 - 26 July 2018
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline 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:
22 March 1996
Deviations:
yes
Remarks:
Actual test item concentration in the high-dose formulation in Week 5 was found to deviate > 15% from nominal (+19.8%). This, and other (minor procedural) deviations were considered not to have compromised the validity or integrity of the study.
GLP compliance:
yes (incl. QA statement)
Limit test:
no
Specific details on test material used for the study:
Correction factor: None (as this would be close to 1).
Final preparation of a solid: According to CiToxLAB France /Study No. 44494 TSR describing the preparation procedure (homogeneity and stability testing) for a range of concentrations covering the lowest and highest used in this study. The dose formulations containing diytterbium trioxide and prepared at 2 mg/mL and 200 mg/mL in 0.5% w/w carboxymethylcellulose aqueous solution were found to be homogenous and stable after 10 days at room temperature.

FORM AS APPLIED IN THE TEST (if different from that of starting material): suspension in a vehicle
Species:
rat
Strain:
Sprague-Dawley
Details on species / strain selection:
The rat was chosen because it is a rodent species accepted by Regulatory Authorities for this type of study. The Sprague-Dawley strain was selected since background data from previous studies are available at CiToxLAB France. This species and strain of rat are recognised as appropriate for general and reproduction toxicity studies. General and reproduction/developmental historical data for this species (same strain and source) are available. This animal model has proved to be susceptible to the effects of reproductive toxicants.
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: 90 rats (42 males and 48 females), Sprague-Dawley, RjHan: SD (Rats CD®) (SPF quality); Janvier, Le Genest-Saint-Isle, France
- Females (if applicable) nulliparous and non-pregnant: yes
- Age at study initiation: (P) males approximately 13 weeks old, females approximately 14 weeks old
- Weight at study initiation: (P) males: mean body weight of 514 g (range: 466 g to 567 g), females: mean body weight of 302 g (range: 273 g to 331 g).
- Fasting period before study: no data
- Housing: The animals were individually housed, except during pairing and lactation, in polycarbonate cages, (Tecniplast 2154, 940 cm²) with stainless steel lids, containing autoclaved sawdust (SICSA, Alfortville, France). Individual housing was chosen in order not to jeopardise gestation and to avoid aggressive behaviour between males around the time of mating. Towards the end of gestation and during lactation, the females and their litters were provided with autoclaved wood shavings (SICSA, Alfortville, France) as nesting material. Each cage contained a nylabone and a rat hut for the environmental enrichment of the animals. The cages were placed in numerical order on the racks.
- Diet (e.g. ad libitum): Free access to SSNIFF R/M-H pelleted maintenance diet, batch No. 89416605 and 64619633 (SSNIFF Spezialdiäten GmbH, Soest, Germany), which was distributed weekly.
- Water (e.g. ad libitum): Free access to bottles containing tap water (filtered with a 0.22 μm filter)
- Acclimation period: Males were acclimated to the study conditions for a period of 29 days before treatment. Females were acclimated to the study conditions for a period of 5 days before the beginning of estrous cycle monitoring during the pre-treatment period. Two supplementary males in the study and two supplementary females per group were acclimated to permit the selection and/or replacement of individuals.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 2°C
- Humidity (%): 50 ± 20%
- Air changes (per hr): about 8 to 15 cycles/hour of filtered, non-recycled air
- Photoperiod (hrs dark / hrs light): 12h/12h

IN-LIFE DATES: From: 2017-07-06 (start of pre-treatment period in females) To: 2017-10-18 (necropsy of last female)
Route of administration:
oral: gavage
Details on route of administration:
The oral route was selected as it is a possible route of human exposure during manufacture, handling or use of the test item. The oral route is a mode of administration recommended by the Regulatory Authorities for this type of study. The dose formulations were administered by oral gavage using a plastic syringe fitted with a plastic gavage tube.
Vehicle:
CMC (carboxymethyl cellulose)
Remarks:
0.5% aqueous solution
Details on oral exposure:
PREPARATION OF DOSING SOLUTIONS
- In the dose formulation study, the dose formulations containing ytterbium oxide prepared at 2 mg/mL and 200 mg/mL in 0.5% CMC were found to be homogenous and stable after 10 days at room temperature. Therefore dose formulation preparation was every 10 days.
- The dose formulations were maintained under delivery conditions (at room temperature, protected from light) throughout the administration procedure.
- Storage condition of control dose formulation: at room temperature, protected from light
- The control dose formulation was stirred just before administration and the test item dose formulations were re-homogenised under magnetic stirring for at least 45 minutes before administration. Before starting the re-homogenisation by magnetic stirring, a spatula dedicated to each test item dose formulation was used to scrape the test item from the bottom of the formulation flasks.
- The formulations were maintained under continuous magnetic stirring throughout the dosing procedure.

VEHICLE
- Justification for use and choice of vehicle (if other than water): based on trial formulations performed by CiToxLAB France, Study No. 44494 TSR
- Concentration in vehicle: 0 (0 mg/kg bw/day); 22 mg/mL (110 mg/kg bw/day); 66 mg/mL (330 mg/kg bw/day); 200 mg/mL (1000 mg/kg bw/day)
- Amount of vehicle (if gavage): 5 mL/kg/day
- Lot/batch no.: SLBQ8545V
- Purity: no data
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Analytical technique: Inductively Coupled Plasma Mass Spectrometry detection (ICP-MS)
Principle and validation of the method: Analytical method provided by the Sponsor and validated at CiToxLAB France (CiToxLAB France/Study No. 44493 VAS) prior to dose formulation analysis.

Determination of test item concentrations in dose formulations:
- Twice in the study (i.e. once in Weeks 1 and 5 of treatment) and in Week 6 for the high dose group.
- A sample was taken from control and test item dose formulations from all dose groups and analysed using the validated method.

Acceptance criterion:
- Measured concentration = nominal concentration ± 15% (85-115%).

Results:
- The test item concentrations in the analysed dose formulations during the study (i.e. in Weeks 1, 5 and 6) remained within an acceptable range of variation (+1.6% to +13.9%) compared to the nominal values (nominal concentrations +/-15% required), except for the high-dose formulation in Week 5 for which the result was found to be +19.8% (i.e. concentration: 239.6 mg/mL and dose level: 1198 mg/kg bw/day).
Duration of treatment / exposure:
Males: for 2 weeks before mating, during the mating period (slightly less than 2 weeks), until euthanasia (at least 6 weeks in total)
Females: for 2 weeks before mating, during the mating period (slightly less than 2 weeks), during gestation, during lactation until day 13 p.p. inclusive, or until euthanasia for females that did not deliver
Frequency of treatment:
Once a day, 7 days per week, at approximately the same time of the day, with a maximum of 3 hours and 39 minutes between the earliest and latest administration.
Dose / conc.:
0 mg/kg bw/day (actual dose received)
Remarks:
control group, group 1
Dose / conc.:
110 mg/kg bw/day (actual dose received)
Remarks:
group 2
Dose / conc.:
330 mg/kg bw/day (actual dose received)
Remarks:
group 3
Dose / conc.:
1 000 mg/kg bw/day (actual dose received)
Remarks:
group 4; deviating dose of 1198 mg/kg bw/day observed at the Week 5 analysis
No. of animals per sex per dose:
10 animals/sex/dose; 4 groups
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: The dose levels were selected in agreement with the Sponsor, based on the results of a previous 2-week dose range finding study (CiToxLAB France/Study No. 44494 TSR) performed in the same species. In that study, three groups of Sprague-Dawley rats received the test item daily, by oral administration (gavage) at dose levels of 110, 330 or 1000 mg/kg bw/day for 2 weeks. Another group received the vehicle control, 0.5% (w/w) carboxymethylcellulose aqueous solution, under the same experimental conditions. The dosage volume was 5 mL/kg. As no relevant findings were observed during the study, the same dose levels were used for the present study.

- Rationale for animal assignment (if not random): During the acclimation period, the required number of animals (40 males and 40 females) was selected according to body weight and clinical condition. The animals were allocated to groups (by sex) using a stratified procedure based on body weight (these data are not presented in the report), so that the average body weight of each group was similar. In addition, only females with regular estrous cycles were allocated to the groups (regularity of estrous cycles was confirmed 1 to 3 days before the beginning of the treatment period).
Positive control:
no
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: From arrival, each animal was observed once a day as part of routine examinations. From the start of the treatment period, each animal was observed once a day (i.e., during dose formulation administration), until the day of necropsy, at approximately the same time, for the recording of clinical signs.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: once before the beginning of the treatment period and then once a week until the end of the study (day of necropsy)
- Detailed clinical examinations were performed on all animals
- Observations included (but were not limited to) changes in the skin, fur, eyes, mucous membranes, occurrence of secretions and excretions and autonomic activity (e.g. lacrimation, piloerection, pupil size, unusual respiratory pattern). Changes in gait, posture and response to handling as well as the presence of clonic or tonic movements, stereotypes (e.g. excessive grooming, repetitive circling) or bizarre behaviour (e.g. self mutilation, walking backwards) were also recorded. The day of onset and disappearance of any observed sign was checked.

BODY WEIGHT: Yes
- Time schedule for examinations: males: on the first day of treatment (day 1), then once a week until sacrifice; females: on the first day of treatment (day 1), then once a week until mated, on days 0, 7, 14 and 20 p.c. (post-coitum) (and on the day of euthanasia for females which did not deliver), and on days 1, 4, 8 and 13 p.p.

FOOD CONSUMPTION: Yes
- The quantity of food consumed by each male was measured once a week, from the first day of treatment until the start of the mating period.
- The quantity of food consumed by each female was measured once a week, from the first day of treatment until the start of the mating period, during gestation for the intervals Days 0-7, 7-14 and 14-20 p.c. and during lactation for the intervals Days 1-4, 4-8 and 8-13 p.p.
- During the mating period, food consumption was not measured for males or females.
- Food intake per animal and per day was calculated by noting the difference between the food given and that in the food-hopper the next time.

FOOD EFFICIENCY: No

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

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes
- Time schedule for collection of blood: the first five males on the day of euthanasia and the first five females euthanised on Day 14 p.p. (all groups)
- Blood samples were collected from the orbital sinus of each animal into tubes containing the appropriate anticoagulant between 8:38 a.m. and 9:33 a.m.
- Anaesthetic used for blood collection: yes, light isoflurane anesthesia
- Animals fasted: yes, prior to blood sampling, the animals were deprived of food for an overnight period of at least 14 hours
- How many animals: the first five males and the first five females (all groups)
- Parameters examined: erythrocytes (RBC), mean cell volume (MCV), packed cell volume (PCV), hemoglobin (HB), mean cell hemoglobin concentration (MCHC), mean cell hemoglobin (MCH), thrombocytes (PLT), leucocytes (WBC), differential white cell count with cell morphology (neutrophils (N), eosinophils (E), basophils (B), lymphocytes (L), large unstained cells (LUC), monocytes (M), reticulocytes (RTC), blood coagulation parameters: prothrombin time (PT), fibrinogen (FIB), activated partial thromboplastin time (APTT)

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: the first five males on the day of euthanasia and the first five females euthanised on Day 14 p.p. (all groups)
- Blood samples were collected from the retro-orbital sinus of each animal into tubes containing the appropriate anticoagulant between 8 a.m. and 11 a.m.
- Animals fasted: yes, prior to blood sampling, the animals were deprived of food for an overnight period of at least 14 hours
- How many animals: the first five males and the first five females (all groups)
- Parameters examined: sodium (Na+), potassium (K+), chloride (Cl-), calcium (Ca++), inorganic phosphorus (PHOS), glucose (GLUC), urea (UREA), creatinine (CREAT), total bilirubin (TOT.BIL), total cholesterol (CHOL), triglycerides (TRIG), alkaline phosphatase (ALP), alanine, aminotransferase (ALAT), aspartate, aminotransferase (ASAT), total proteins (PROT), albumin (ALB), albumin/globulin ratio (A/G), bile acids (BIL.AC).

THYROID HORMONES: Yes
- Blood samples were taken in the morning (between 7:30 and 10 a.m.) into tubes containing K3-EDTA as anticoagulant.
- Blood samples were taken at termination on Day 14 p.p. from all F0 females and at termination from all F0 males (approximately 0.5 mL of blood was collected from the orbital sinus under isoflurane anaesthesia).
- Blood was centrifuged within 2 hours after sampling (approximately 3000g for 10 minutes at +4°C). The plasma was transferred into two separate tubes (when possible at least 125 μL in the first tube and the remaining plasma in the second tube) and frozen at -80°C.
- The levels of the thyroid hormone (T4) and thyroid stimulating hormone (TSH) were determined by Luminex MAP® technology for F0 males sampled at termination.
- Plasma samples obtained on Day 14 p.p. from F0 females are kept at -80°C. In agreement with the Sponsor, no analyses were performed for these animals (as there was no indication of an effect on the thyroid glands).

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: Yes
- Time schedule for examinations: once over a 60-minute period
- Battery of functions tested: motor activity

IMMUNOLOGY: No

FUNCTIONAL OBSERVATIONAL BATTERY: Yes
- The first five males and the first five surviving females euthanised on Day 14 p.p. from each group were evaluated with a Functional Observation Battery once at the end of the treatment period. For females, this was performed on Day 13 p.p. after euthanasia of the pups.
- All animals were observed in the cage, in the hand and in the standard arena.
- Detailed clinical examinations: the following parameters were assessed and graded: in the cage: touch escape or ease of removal from the cage; in the hand: fur appearance, salivation, lacrimation, piloerection, exophthalmos, reactivity to handling, pupil size (presence of myosis or mydriasis); in the standard arena (2-minute recording): grooming, palpebral closure, defecation, urination, tremors, twitches, tonic and clonic convulsions, gait, arousal (hypo- and hyper-activity), posture, stereotypy, behaviour, breathing, ataxia and hypotonia
- Reactivity to manipulation and different stimuli: the following measurements, reflexes and responses were recorded: touch response, forelimb grip strength, pupillary reflex, visual stimulus response, auditory startle reflex, tail pinch response, righting reflex, landing foot splay, at the end of observation: rectal temperature

MORTALITY AND MORBIDITY: Yes
- Each animal was checked for mortality and morbidity once a day before the treatment period and at least twice a day (early in the morning and close to the end of the working day) during the treatment period, including weekends and public holidays. Attention was paid to humane end-points.
Sacrifice and pathology:
SACRIFICE
- Male animals: On completion of the treatment period, after the end of the mating period (at least 6 weeks of treatment in total), after at least 14 hours (maximum 24 hours) fasting (with water available), all surviving F0 animals were deeply anesthetised by an intraperitoneal injection of sodium pentobarbital and euthanised by exsanguination.
- Maternal animals: On completion of the treatment period, on day 14 p.p., after at least 14 hours (maximum 24 hours) fasting (with water available), all surviving F0 animals were deeply anesthetised by an intraperitoneal injection of sodium pentobarbital and euthanised by exsanguination. The following F0 females were euthanised by CO2 inhalation followed by cervical dislocation without overnight fasting: female K28872 (group 1), female K28877 (group 2) and females K28899 and K28902 (group 4) which did not deliver: on day 26 p.c. (after a body weight recording to check for a possible un-noticed delivery); female K28862 (group 1) and female K28886 (group 3) which had difficulties to deliver: on day 24 p.c. (after a body weight recording).

GROSS PATHOLOGY
- Gross necropsy consisted of external and internal examinations including the cervical, thoracic, and abdominal viscera.
- A complete macroscopic post-mortem examination was performed on all F0 animals including the females euthanised prematurely. This included examination of the external surfaces, all orifices, the cranial cavity, the external surfaces of the brain and spinal cord, the thoracic, abdominal and pelvic cavities with their associated organs and tissues and the neck with its associated organs and tissues. Special attention was paid to the reproductive organs. The numbers of corpora lutea and implantation sites were recorded for females euthanised as scheduled on Day 14 p.p. For prematurely euthanised females, pregnancy status was determined and the numbers of corpora lutea and implantation sites were recorded and classified as live or dead concepti, early or late resorptions or scars.

HISTOPATHOLOGY
- The body weight of each animal euthanised as scheduled (after the end of the mating period for males or on Day 14 p.p. for females) was recorded before euthanasia.
- The following organs of the first 5 euthanised-as-scheduled males and the first 5 females euthanised on Day 14 p.p. of each group were weighed wet as soon as possible after dissection: adrenals, brain (including medulla/pons, cerebellar and cerebral cortex), epididymides, heart, kidneys, liver, pituitary gland, prostate (dorso-lateral and ventral), seminal vesicles (including coagulating glands), spleen, testes, thymus, thyroids with parathyroids. The following organs were weighed for all animals euthanised as scheduled: epididymides, prostate (dorso-lateral and ventral), seminal vesicles (including coagulating glands), testes, thyroids with parathyroids.
- The ratio of organ weight to body weight (recorded immediately before euthanasia) was calculated.
- The following tissues from the first 5 euthanised-as-scheduled males and the first 5 females euthanised on Day 14 p.p., and, additionally, all animals prematurely euthanised, were preserved in 10% buffered formalin (except for the eyes with optic nerves and Harderian glands, and the testes and epididymides which were fixed in modified Davidson's fixative): macroscopic lesions (all animals), adrenals, brain (including medulla/pons, cerebellar and cerebral cortex), cecum, colon, Cowper's (i.e. bulbo-urethral) glands (all animals), duodenum, epididymides (all animals), esophagus, eyes with Harderian glands, femoral bone with articulation, glans penis (all animals), gut-associated lymphoid tissue (GALT), heart, ileum, jejunum, kidneys, levator ani plus bulbo cavernous muscle complex (all animals), liver, lungs with bronchi, lymph nodes (mandibular and mesenteric), mammary gland area (all animals), ovaries (with oviducts) (all animals), pituitary gland, prostate (dorso-lateral and ventral) (all animals), rectum, sciiatic nerve, seminal vesicles (including coagulating glands) (all animals), skeletal muscle, spinal cord (cervical, thoracic and lumbar), spleen, sternum with bone marrow, stomach with forestomach, testes (all animals), thymus, thyroids with parathyroids (all animals), trachea, urinary bladder, uterus (horns and cervix) (all animals), vagina (all animals).
- All tissues required for microscopic examination were trimmed based on the RITA guidelines, when applicable, embedded in paraffin wax, sectioned at a thickness of approximately 4 microns and stained with hematoxylin-eosin (except testes and epididymides which were stained with hematoxylin/PAS). This tissue processing was performed at CiToxLAB France.
- A microscopic examination was performed on: all tissues listed above from the first five euthanised-as-scheduled males and the first five females euthanised on Day 14 p.p. of the control and high dose groups (groups 1 and 4); all macroscopic lesions of all groups; reproductive organs from any animal that did not conceive or from pregnant females that did not deliver, to investigate possible causes (this also includes group 4 male K28396).
- Special emphasis was paid to the stages of spermatogenesis in the male gonads and histopathology of interstitial testicular cell structure.
Other examinations:
No further data
Statistics:
Body weight, food consumption and reproductive data:
- Data were compared by one-way analysis of variances and Dunnett test (mean values being considered as normally distributed, variances being considered as homogenous) or by Fisher’s exact probability test (proportions).

Hematology and blood biochemistry:
- A sequence was used for the statistical analyses of hematology and blood biochemistry data. According to the sequence, the Dunn test, Dunnett test or Mann-Whitney/Wilcoxon test were used to analyse the data.

Organ weight:
- PathData software was used to perform the statistical analysis of organ weight data (level of significance: 0.05 or 0.01) according to a sequence. At the end of the sequence, the data were analysed using either Dunn test or Dunnett test.
Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
Main clinical signs in animals that were not prematurely euthanised due to delivery difficulties:
- At 1000 mg/kg bw/day, whitish coloured feces was observed from Day 28 in 8/10 males and from Day 8 p.c. at the earliest in all females. This finding was considered to result from the whitish colour of the dose formulation. The relevance of this finding remains undetermined.
- Piloerection, associated with pallor of extremities, was noted for 1 or 2 days at the beginning of the lactation period in 1/9 females (female K28888) given 330 mg/kg bw/day. These findings were also seen in 1/8 females (female K28898) given 1000 mg/kg bw/day along with round back, soiled urogenital area, reddish coloured urine and brownish vaginal discharge for up to 2 days after delivery. As these clinical signs were recorded with a similar incidence in controls at delivery (i.e. control female K28862 prematurely euthanised on Day 24 p.c. due to difficulties to deliver), a test item relationship was considered to be unlikely.
- All other clinical signs (i.e. chromorhinorrhea, chromodacryorrhea, areas of hair loss, cutaneous lesions, scabs and/or short/broken teeth) were considered to be unrelated to the test item treatment as they were present in control animals, not dose-related, reported sporadically in only a few animals and/or commonly observed findings in this species and strain.

Clinical findings in prematurely euthanised females (observations prior to euthanasia):
- Female K28862 (control group, euthanised on Day 24 p.c. due to difficulties to deliver): piloerection, round back, pallor of extremities, dyspnea, soiled urogenital area and reddish coloured urine.
- Female K28872 (control group, euthanised on Day 26 p.c. for no delivery): no relevant clinical signs were observed during premating or gestation periods.
- Female K28877 (110 mg/kg bw/day, euthanised on Day 26 p.c. for no delivery): no relevant clinical signs were observed during premating or gestation periods.
- Female K28886 (330 mg/kg bw/day, euthanised on Day 24 p.c. due to difficulties to deliver): piloerection, round back, pallor of extremities and eyes, coldness to the touch, dyspnea, soiled urogenital area and reddish vaginal discharge. In the absence of similar observations at 1000 mg/kg bw/day, any relationship of these observations to the test item was excluded. Difficulties to deliver were moreover noted with the same incidence in the control group and were therefore considered to be spontaneous and therefore unrelated to the test item.
- Females K28899 and K28902 (1000 mg/kg bw/day, euthanised on Day 26 p.c. for no delivery): no relevant clinical signs were observed during the premating period and whitish coloured feces was observed from Day 9 or 12 p.c.
Mortality:
no mortality observed
Description (incidence):
There were no unscheduled deaths in males.
In females, there were two unscheduled sacrifices (due to difficulties to deliver).
Body weight and weight changes:
no effects observed
Description (incidence and severity):
- There were no effects on mean body weight or mean body weight change.
- Statistically significant differences between control and test item-treated animals consisting of higher mean body weight gain in males given 1000 mg/kg bw/day between Days 8 and 15 and lower mean body weight gain in males given 330 mg/kg bw/day between Days 15 and 22, and very slight mean body weight loss in females given 110 or 330 mg/kg bw/day (-1 to -3 g vs. +3 g in controls between Days 1 and 8 of the premating period). These findings were considered to be of no toxicological importance as they were isolated, of slight magnitude and/or not dose-related.
Food consumption and compound intake (if feeding study):
no effects observed
Description (incidence and severity):
There were no effects on mean food consumption.
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
effects observed, non-treatment-related
Description (incidence and severity):
- At 1000 mg/kg bw/day, statistically significant lower mean white blood cell count was noted in males (-19% vs. controls, p<0.05). In the absence of any other significant changes, and as values remained within the Historical Control Data range, this difference was considered to be of no toxicological importance.
- There were no effects on hematological parameters at any other dose level.
Clinical biochemistry findings:
effects observed, non-treatment-related
Description (incidence and severity):
- Statistically significant lower mean urea levels were noted in males from 110 mg/kg bw/day.
- These differences were not dose-related, were not associated with other blood biochemistry or histological changes and were within the range of the HCD, they were therefore considered to be of no toxicological importance.
- All the other differences (namely lower mean sodium levels in low- and intermediate-dose males and lower mean calcium levels in low- and high-dose males) were slight and not dose-related. Therefore, they were considered to be of no toxicological importance.
- The thyroid hormone levels were considered to be unaffected by the test item treatment in F0 males.
- Higher mean T4 concentration in males given 1000 mg/kg bw/day and lower mean TSH concentrations in males given 110 or 1000 mg/kg bw/day were considered to be of no toxicological significance in the absence of statistical significance and correlating effects at pathology, and in view of the slight differences from control means.
- At 330 mg/kg bw/day and when compared with controls, there was a statistically significant decrease in mean TSH concentration (-45%). This observation was considered as fortuitous in the absence of a dose relationship.
Urinalysis findings:
not examined
Behaviour (functional findings):
no effects observed
Description (incidence and severity):
- There were no test item treatment-related neurologic abnormalities.
- An excessive quantity/abnormal appearance of urine was noted in 1/5 males given 1000 mg/kg bw/day. Higher mean landing foot splay values were noted in males given 110 or 1000 mg/kg bw/day (89 and 83 mm vs. 69 mm in controls, respectively).
- These findings were considered to be of no toxicological importance as they were poorly dose-related, did not correlate with any other findings and/or were of isolated occurrence.
- There were no differences between test item-treated and control groups in motor activity (horizontal movements and rearing).
- The lower mean number of horizontal movements noted in females given 110 mg/kg bw/day was not dose related, and no correlation with a lower mean number of rearing movements was recorded. Therefore this was not considered to be test item treatment related.
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
effects observed, non-treatment-related
Description (incidence and severity):
- There were no organ weight changes related to administration of the test item at the end of the treatment period.
- The absolute and relative adrenal weights were higher in females treated at 330 or 1000 mg/kg bw/day without reaching a statistical significance. As this was mainly due to individual variations, any relationship to the test item was considered to be unlikely.
- The absolute and relative thymus weights were higher in females treated at 1000 mg/kg bw/day without reaching a statistical significance. In the absence of histopathological correlates any relationship to the test item was considered to be unlikely.
- The absolute and relative seminal vesicle weights were statistically significantly higher at 110 mg/kg bw/day (p<0.05) and the relative spleen weight lower in males at 330 mg/kg bw/day. In the absence of a similar trend at higher dose levels, any relationship to the test item was excluded.
- Other changes in the mean organ weights were considered to be part of the normal individual variation and without any relationship to the test item.
Gross pathological findings:
effects observed, non-treatment-related
Description (incidence and severity):
- There were no test item-related changes observed at necropsy.
- The right testes of male K28391 treated at 1000 mg/kg bw/day was small (recorded as reduced) correlating histologically with marked atrophy degeneration of tubules. As this observation was unilateral, observed in one male only at this dose level and occasionally seen in rats (Mc Innes, 2012), any relationship to the test item was excluded.
- Agenesis of the bulbourethral glands was observed in male K28399 treated at 1000 mg/kg bw/day. This isolated observation was considered to be incidental.
- The above described findings did not affect the mating or fertility index as the corresponding females were pregnant.
- Control female K28862 (euthanised on Day 24 p.c. due to difficulties to deliver) showed red discoloration in the vagina at necropsy. In the uterus, dead fetuses were observed in both horns.
- Female K28886 (330 mg/kg bw/day, euthanised on Day 24 p.c. due to difficulties to deliver) had translucent fluid in the thoracic cavity and the thymus was gelatinous. A tan discoloration of mandibular glands and mandibular lymph nodes, liver and kidneys was observed, correlating histologically with necrosis in the liver and kidneys. Scars and placenta were observed in the uterus of this animal.
- Abovementioned findings in the females euthanised on Day 24 p.c. due to difficulties to deliver were not considered to be related to treatment with the test item.
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
effects observed, non-treatment-related
Description (incidence and severity):
- There were no test item-related changes at microscopic examination in terminal sacrificed animals.
- Changes observed were considered to be either incidental or part of the normal background commonly seen in rats and without any relationship to the test item.
- Erosion of the stomach was observed in one control female and one female treated at 1000 mg/kg bw/day which had also slight erosion in the cecum. Erosion of the stomach is commonly seen in rats (Mc Innes, 2012) and considered to be stress-related. Erosions in the cecum are not commonly seen in rats, therefore, the cecal erosion in a single female at 1000 mg/kg bw/day was considered to be of doubtful toxicological significance.
- Female K28886 treated at 330 mg/kg bw/day had marked acute tubular necrosis in the renal cortex. Eosinophilic material, consistent with fibrin, was also seen in capillaries of the glomeruli. In the liver, slight multifocal centrilobular hepatocellular necrosis was observed. The origin of these degenerative changes remained unclear, however, a relationship to difficulties of this animal to deliver is possible. In the absence of similar changes at 1000 mg/kg bw/day any relationship of these lesions to the test item was excluded. Additional changes in this animal consisted of interstitial edema in the thymus which correlated with necropsy change and slight to moderate increased apoptotic cell numbers in the thymus and mandibular lymph nodes which were considered to be non-specific (stress related).
- Slight multifocal macrophage aggregates were observed in lungs of female K28903 treated at 1000 mg/kg bw/day. In the cytoplasm of these macrophages, there were birefringent grey crystals which may correspond to the test item. Aspiration of the test item during the gavage was considered to be the most likely cause of this observation.
- Careful examination of testes and other male genital organs did not show any test item-related changes, particularly for males K28393 and K28396 treated at 1000 mg/kg bw/day for which corresponding females were not pregnant.
- There were no test item-related histopathological changes in the reproductive organs of females K28889 and K28902 treated at 1000 mg/kg bw/day which could explain the absence of pregnancy in these animals.
- Other changes observed at the histopathological examination were considered to be part of the normal background commonly seen in rats.
Histopathological findings: neoplastic:
not examined
Other effects:
not specified
Details on results:
No further data
Key result
Dose descriptor:
NOAEL
Remarks:
for parental systemic toxicity
Effect level:
>= 1 000 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Remarks on result:
not determinable due to absence of adverse toxic effects
Key result
Critical effects observed:
no
Conclusions:
Interpretation of results: GHS criteria not met

The test item, diytterbium trioxide, was administered daily by oral gavage to male and female Sprague Dawley rats, for 2 weeks before mating, during mating and (for females) throughout gestation and until Day 13 post-partum, at the dose-levels of 110, 330 or 1000 mg/kg bw/day.
Based on the experimental conditions of the study the No Observed Adverse Effect Level (NOAEL) was considered to be higher than or equal to 1000 mg/kg bw/day based on the absence of adverse findings (i.e. findings of toxicological relevance) related to treatment with the test item at this high dose level.
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
1 000 mg/kg bw/day
Study duration:
subacute
Species:
rat

Repeated dose toxicity: inhalation - systemic effects

Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: inhalation - local effects

Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: dermal - systemic effects

Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: dermal - local effects

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

Repeated dose toxicity - oral

In a reliable, GLP-compliant study performed according to OECD guideline 422 (Papineau, 2018), the test item was administered once daily to 3 groups of 10 male and 10 female Sprague-Dawley rats by oral administration (gavage) at dose levels of 110, 330 or 1000 mg/kg bw/day. The dosing period started 2 weeks before mating, continued during mating, and for females, throughout gestation until day 13 post-partum. The test item was administered in the vehicle (0.5% carboxymethylcellulose aqueous solution) under a constant dosage volume of 5 mL/kg bw/day. One other group of 10 males and 10 females received the vehicle alone and acted as a control group.

Actual concentrations of the test item in the dose formulations analysed during the study remained within an acceptable range of variations (+1.6% to +13.9% - acceptable is +/-15% from nominal values) except for the high dose formulation in week 5, for which the result was found to be +19.8% (corresponding to an actual dose level of 1198 mg/kg bw/day) when compared to nominal values. This deviation was not considered to have affected the integrity or interpretation of the results.

No test-item related unscheduled deaths were observed during the study. No test item-related clinical signs were observed apart from whitish coloured feces in all males at 1000 mg/kg bw/day and also appearing in all females during the gestation period at this dose level. As these findings resulted from the whitish colour of the dose formulations, they were considered related to the test item, but not relevant from a toxicological point of view. No effects were observed at any dose level on the outcome of functional observation battery tests or motor activity data, body weight or body weight change, and food consumption. Concerning hematology and blood biochemistry, all observed changes were considered to be of no toxicological importance as they were isolated, poorly dose-related, and/or within the range of the historical control data. There were no test item-related effects on the thyroid hormone levels in F0 males (females not studied) at any dose level. Finally, there were no test item-related changes in mean organ weights or at macroscopic or microscopic examination. Based on the observations made during this study, the NOAEL for parental systemic toxicity was considered to be higher than or equal to 1000 mg/kg bw/day, since there were no adverse findings related to the test item at this highest dose level. The study was considered as the key study for endpoint coverage.

Repeated dose toxicity - inhalation/dermal

No key studies were identified for repeated dose toxicity after inhalation or dermal exposure.

A key study is available for the oral route of exposure. According to the REACH Regulation, only one route of exposure should be tested for repeated dose toxicity (Column 2, Annex VIII, Section 8.6.1). Therefore, it is not necessary to perform a repeated dose toxicity study via the inhalation or dermal route of exposure.

Justification for classification or non-classification

Repeated dose toxicity - oral

Based on the NOAEL value of higher than or equal to 1000 mg/kg bw/day, the test substance is not to be classified for Specific Target Organ Toxicity after Repeated Exposure (STOT RE) according to the CLP Regulation.

Repeated dose toxicity - inhalation

No key repeated dose toxicity study via inhalation is available.

Repeated dose toxicity - dermal

No key repeated dose toxicity study via dermal application is available.