Hexakis[μ-(acetato-O:O')]-μ3-oxo-triangulo-triruthenium acetate

Repeated dose toxicity: oral

Administrative data

Endpoint:

short-term repeated dose toxicity: oral

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

"during 2004"

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Remarks:

Full study report not available. The information provided is limited (particularly with regards to test material identity and methodology) to a summary (TSCA submission) of the draft study report and a subsequent expert apprasial

Data source

Reference

Reference 1

Reference Type:

other company data

Title:

Unnamed

Year:

2015

Report date:

2015

Reference 2

Reference Type:

other company data

Title:

Unnamed

Year:

2005

Report date:

2005

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 407 (Repeated Dose 28-Day Oral Toxicity Study in Rodents)

Version / remarks:

The study design was based on the 1995 version. It included a Functional Observation Battery segment. No satellite (recovery) groups were incorporated.

Deviations:

yes

Remarks:

The dose level intervals exceeded the two- to four-fold steps considered optimal in study design, but did not exceed the typically applied maximum of ten-fold intervals.

GLP compliance:

yes

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Hexakis[μ-(acetato-O:O')]-μ3-oxo-triangulo-triruthenium acetate

EC Number:

259-653-7

EC Name:

Hexakis[μ-(acetato-O:O')]-μ3-oxo-triangulo-triruthenium acetate

Cas Number:

55466-76-7

Molecular formula:

C12H18O13Ru3.C2H3O2

IUPAC Name:

hexamethyl-2λ³-oxa-4λ³-oxa-6λ³-oxa-8λ³-oxa-10λ³-oxa-12λ³-oxa-13λ³-oxa-15λ³-oxa-16λ³-oxa-18λ³-oxa-19λ³-oxa-21λ³,22λ¹-dioxa-1,5,9-triruthenahexacyclo[7.3.3.3¹,⁵.3⁵,⁹.1¹,⁵.0⁹,²²]docosa-3,7,11,14,17,20-hexaene-1,1,1,5,5,5,9,9,9-nonakis(ylium)-2,6,10,13,16,19-hexaide-22,22-diuide acetate

Test material form:

not specified

Details on test material:

not specified

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: not specified
- Expiration date of the lot/batch: not specified
- Purity test date: not specified

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: not specified
- Stability under test conditions: not specified
- Solubility and stability of the test substance in the solvent/vehicle: not specified
- Reactivity of the test substance with the solvent/vehicle of the cell culture medium: not specified

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing: not specified
- Preliminary purification step (if any): not specified
- Final dilution of a dissolved solid, stock liquid or gel: not specified
- Final preparation of a solid: not specified

FORM AS APPLIED IN THE TEST (if different from that of starting material) not specified

Test animals

Species:

rat

Strain:

other: Crl:CD(SD) IGS BR

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Not specified.
- Age at study initiation: Not specified.
- Weight at study initiation: Not specified.
- Fasting period before study: Not specified.
- Housing: Not specified.
- Diet (e.g. ad libitum): Not specified.
- Water (e.g. ad libitum): Not specified.
- Acclimation period: Not specified.


ENVIRONMENTAL CONDITIONS
- Temperature (°C): Not specified.
- Humidity (%): Not specified.
- Air changes (per hr): Not specified.
- Photoperiod (hrs dark / hrs light): Not specified.

IN-LIFE DATES: From: To: Not specified.

Administration / exposure

Route of administration:

oral: gavage

Details on route of administration:

Not specified.

Vehicle:

water

Details on oral exposure:

PREPARATION OF DOSING SOLUTIONS:

VEHICLE
- Concentration in vehicle: Not specified.
- Amount of vehicle (if gavage): Not specified.
- Lot/batch no. (if required): Not specified.
- Purity: Not specified.

"Solubility was apparently complete at the concentrations used for the dosing formulations" and "Analysed concentraitons versus nominals were within acceptable limits".

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Analysed concentrations versus nominals were within acceptable limits.

Duration of treatment / exposure:

Male and female rats were treated for up to 4 weeks. The group of animals receiving 1000 mg/kg bw/day (the highest tested dose) were treated for only 3 weeks, as this dose level "exceeded the maximum tolerated dose."

Frequency of treatment:

Not specified (presumably daily administration).

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

15 mg/kg bw/day (nominal)

Doses / concentrations 2

Dose / conc.:

150 mg/kg bw/day (nominal)

Doses / concentrations 3

Dose / conc.:

1 000 mg/kg bw/day (nominal)

Remarks:

Dose level exceeded the maximum tolerated dose - group prematurely terminated (at week 3)

No. of animals per sex per dose:

5 animals/sex/group, 3 groups, plus a "similarly constituted control group"

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: Not specified. The dose level intervals exceeded the two- to four-fold steps considered optimal in study design, but did not exceed the typically applied maximum of ten-fold intervals.

Examinations

Observations and examinations performed and frequency:

Limited details of this guideline study provided in available summary and apprasial. According to the former, "During the study, clinical condition, detailed physical and arena observations, sensory reactivity, grip strength, motor activity, bodyweight, food consumption, water consumption, haematology, blood chemistry, organ weight, macropathology and histopathology investigations were undertaken".

Sacrifice and pathology:

GROSS PATHOLOGY: Yes

HISTOPATHOLOGY: Yes

Other examinations:

Included a Functional Observation Battery (FOB) segment.

Statistics:

Not specified.

Results and discussion

Results of examinations

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

At 1000 mg/kg bw/day (the highest tested dose), signs observed at detailed physical examination showed a general deterioration in clinical condition with observations such as thin build, respiratory problems, a dark discharge from the anus and dark staining on the tails. Also following anaesthesia for blood sampling the recovery times for these animals was noted to be longer than expected. Cage tray observations revealed green staining and dark faeces. Signs observed in association with dosing were: resistance to insertion of the catheter, salivation and noisy respiration. High dose group prematurely terminated at week 3.

At the low- and mid-dose groups (15 and 150 mg/kg bw/day respectively) no clinical signs were observed except darkened faeces.

Mortality:

mortality observed, treatment-related

Description (incidence):

At 1000 mg/kg bw/day, 1 male and 2 females were killed in extremis.

No other deaths occurred.

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

At 1000 mg/kg bw/day, rats showed slightly low bodyweight gain during the first 2 weeks of treatment, with either bodyweight loss or stasis during the third week of treatment.

Bodyweight gain was unaffected at the low- and mid-dose levels.

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Description (incidence and severity):

At 1000 mg/kg bw/day, "food consumption was low".

Food consumption was unaffected at the low- and mid-dose levels.

Food efficiency:

effects observed, treatment-related

Description (incidence and severity):

At 1000 mg/kg bw/day, the food conversion efficiency was low during the first 2 weeks of treatment and was subsequently incalculable during the third week of treatment.

Food efficiency was unaffected at the low- and mid-dose levels.

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

effects observed, treatment-related

Description (incidence and severity):

At the highest tested dose, water consumption was elevated during the third week of treatment.

Water consumption was unaffected at the low- and mid-dose levels.

Ophthalmological findings:

not specified

Haematological findings:

effects observed, treatment-related

Description (incidence and severity):

At 1000 mg/kg bw/day, white blood cell counts were "unselectively high" and slight/moderate hyperchromasia was apparent. Platelet counts were high, with males showing a decrease in the activated partial thromboplastin clotting time.

At 15 and 150 mg/kg bw/day, late stage elevations in white blood cell counts (including neutrophils) were observed, though these effects were not statistically significant.

"Group mean total white blood cell, neutrophil and lymphocyte counts showed elevations for females in all treated groups in a dose-related manner (although not achieving statistical significance). A similar, though less clear trend, was evident in respect of mean total white blood cell count and neutrophils for males [of the mid- and high-dose groups]."

Clinical biochemistry findings:

effects observed, treatment-related

Description (incidence and severity):

Males receiving 1000 mg/kg bw/day showed high alanine amino-transferase activity and potassium levels. Females of this dose group showed high alanine-amino transferase activity, aspartate amino transferase activity and urea levels.

Males receiving 15 or 150 mg/kg bw/day had low albumin levels, with males at 150 mg/kg bw/day showing low total protein levels. Also, aspartate amino-transferase levels were slightly elevated for females at 15 or 150 mg/kg bw/day.

Urinalysis findings:

not specified

Behaviour (functional findings):

not specified

Immunological findings:

not specified

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Description (incidence and severity):

After 3 weeks of treatment, relative adrenal and spleen weights were high for both sexes, and males showed low thymus weights at the highest tested dose.

Absolute and relative spleen weights in males that received 15 or 150 mg/kg bw/day were high. Females receiving 150 mg/kg bw/day showed low relative heart weights.

Gross pathological findings:

effects observed, treatment-related

Description (incidence and severity):

At 1000 mg/kg bw/day, the majority of macroscopic observations were confined to the gastro-intestinal tract with abnormal content, distension and thickening. Dark areas were observed primarily on the stomach wall and the limiting ridge was also thickened.

At 150 mg/kg bw/day, the "majority of females" showed abnormal gastrointestinal content at macroscopic examination.

No treatment-related effects were seen in the low-dose group.

Neuropathological findings:

effects observed, treatment-related

Description (incidence and severity):

Degenerative peripheral and central nervous system lesions were detected at the top dose, generally described as "minimal" with the exception of two gradings of "slight" in relation to spinal cord lesions. In the sciatic nerve, degernate fibres were noted for all high-dose animals, and there was an increase in the incidence/severity of degernate fibres in the cervical, thoracic and lumbar section of the spinal cord.

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

At 1000 mg/kg bw/day: In the spleen, myeloid hyperplasia (graded "slight" or "minimal") in the red pulp was evident in "the majority of animals", which may suggest extramedullary hematopoiesis as a secondary response to inflammatory changes elsewhere. Examination of the stomach showed ulceration, focal epithelial hyperplasia, subepithelial oedema, and inflammatory cell infiltrate in the nonglandular region, with epithelial necrosis, epithelial hyperplasia, hyperkeratosis and subepithelial oedema at the limiting ridge. Focal erosions, or necrosis, of fundic and/or antral mucosa, mucosal haemorrhage, submucosal oedema, and inflammatory cell infiltrate, were noted in the glandular region of the stomach. Examination of the intestines, showed focal mucosal necrosis, mucosal haemorrhage, and submucosal inflammatory cell infiltrate.


Microscopic examination of the spleen revealed myeloid hyperplasia in the red pulp for 1 male that received 15 mg/kg bw/day, and "a few animals" at 150 mg/kg bw/day.

Histopathological findings: neoplastic:

not examined

Other effects:

not specified

Effect levels

1

Key result

Dose descriptor:

NOAEL

Effect level:

15 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: No significant, treatment-related effects observed at this dose-level.

2

Dose descriptor:

LOAEL

Effect level:

150 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

histopathology: non-neoplastic

organ weights and organ / body weight ratios

Target system / organ toxicity

Key result

Critical effects observed:

yes

Lowest effective dose / conc.:

150 mg/kg bw/day (nominal)

System:

haematopoietic

Organ:

leucocyte development

spleen

Treatment related:

yes

Dose response relationship:

yes

Relevant for humans:

yes

Applicant's summary and conclusion

Conclusions:

In a 4-week repeated dose toxicitystudy, to GLP, ruthenium acetate was administered by gavage to CD rats at 15, 150 or 1000 mg/kg bw/day. Effects at 1000 mg/kg bw/day exceeded the maximum tolerated dose and the study was terminated prematurely at week 3. The lower doses were generally well tolerated with findings restricted to minor variations in clinical pathology parameters and effects on the spleen. Although considered of low-severity, the effects on the spleen (enlargement in males and histopathological change) increased in incidence affecting one, to "several", to the "majority", of animals in the low-, mid- and high-dose groups, respectively. The low dose of 15 mg/kg bw/day is therefore considered the most appropriate to assign as the study NOAEL.

Executive summary:

In a repeated dose toxicity study, generally conducted in conformance with OECD Test Guideline 407 and performed to GLP, CD rats (5/sex/group) were administered ruthenium acetate (CAS RN 55466-76-7) by oral gavage for 4 weeks at dose levels of 15, 150 or 1000 mg/kg bw/day. A control group received vehicle only (water). During the study, clinical condition, detailed physical and arena observations, sensory reactivity, grip strength, motor activity, bodyweight, food consumption, water consumption, haematology, blood chemistry, organ weight, macropathology and histopathology investigations were undertaken, and a Functional Observation Battery (FOB) segment was included.

Administration of 1000 mg/kg bw/day (the highest tesed dose) exceeded the maximum tolerated dose, resulting in mortality, significant clincal signs, effects on haematology and clincial chemistry (indicative of effects on the liver and kidney), and macroscopic and histopathological changes in the spleen, stomach and intestines, and spinal cord and sciatic nerve. The premature termination of the high dose group occurred after 3 weeks of treatment. Doses of 15 or 150 mg/kg bw/day (for 4 weeks) were generally well tolerated with findings restricted to minor variations in clinical pathology parameters and myeloid hyperplasia in the red pulp of the spleen. Although of low-severity, the effects on the spleen (enlargement in males and coincident histopathological change) increased in incidence affecting one, to "several", to the "majority", of animals in the low-, mid- and high-dose groups, respectively. As such, the low dose group (15 mg/kg bw/day) appears the most appropriate to assign as a robust and health-precautionary study NOAEL.