Ruthenium trichloride hydrate

Repeated dose toxicity: oral

Administrative data

Endpoint:

short-term repeated dose toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

14 July 2016-28 February 2017

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

Study conducted according to GLP

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2017

Report date:

2017

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

03 October 2008

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Ruthenium (III) chloride hydrate

IUPAC Name:

Ruthenium (III) chloride hydrate

Test material form:

solid: particulate/powder

Test animals

Species:

rat

Strain:

Wistar

Details on species / strain selection:

Wistar rat as a rodent is one of the standard strains for repeat-dose toxicity studies.

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories, Research Models and Services, Germany GmbH, Sandhofer Weg 7, D-97633, Sulzfeld, Germany.
- Females (if applicable) nulliparous and non-pregnant: Yes.
- Age at study initiation: Approximately 8 weeks old at onset of treatment.
- Weight at study initiation: Males: 283-347 g. Females: 191-249 g. Weight did not exceed +/- 20% of mean weight at onset of treatment.
- Fasting period before study: Not specified.
- Housing: Type II polycarbonate/ wire grid bottomed cages. Animals were housed two or three per cage.
- Diet (e.g. ad libitum): ssniff® SM R/M-Z+H “Complete Diet for Rats and Mice- Breeding and Maintenance” (ssniff Spezialdiäten GmbH, D-59494 Soest Germany), with or without Ruthenium chloride (treated or control groups, respectively) provided ad libitum.
- Water (e.g. ad libitum): Tap water provided ad libitum.
- Acclimation period: 15 days.

DETAILS OF FOOD AND WATER QUALITY: Tap water as for human consumption, assessed once every three months, with monthly microbiological assessment.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20.3 - 27.4 °C
- Humidity (%): 33 - 78%
- Air changes (per hr): 15-20.
- Photoperiod (hrs dark / hrs light): 12 hours artificial light (6am - 6pm). 12 hours dark.

IN-LIFE DATES: From: 30 June 2016 (animal arrival) To: 25 August 2016 (last necropsy).

Administration / exposure

Route of administration:

oral: feed

Details on oral exposure:

DIET PREPARATION
- Rate of preparation of diet (frequency): A single diet mix batch was prepared before the study.
- Mixing appropriate amounts with (Type of food): About 6 minutes for pre-mixing and 4-8 minutes for diet mixing.
- Storage temperature of food: approximately 15-21 °C (storage areas); 22 +/- 3 °C (animal rooms).

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Analysis of the diets for homogeneity and concentration of Ruthenium chloride were performed at the Test Site using a validated ICP/AES method to determine the Ruthenium content.

Duration of treatment / exposure:

28 days.

Frequency of treatment:

Constant (dietary).

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

0 ppm

Remarks:

Control, received diet prepared in a similar manner to treatment groups, without inclusion of test material.

Doses / concentrations 2

Dose / conc.:

500 ppm

Remarks:

"Low dose" group. Approximately 40.6 mg/kg bw/day.

Doses / concentrations 3

Dose / conc.:

1 500 ppm

Remarks:

"Mid dose" group. Approximately 124 mg/kg bw/day.

Doses / concentrations 4

Dose / conc.:

5 000 ppm

Remarks:

"Mid high dose" group. Approximately 407 mg/kg bw/day.

Doses / concentrations 5

Dose / conc.:

15 000 ppm

Remarks:

"High dose" group. Approximately 1238 mg/kg bw/day.

No. of animals per sex per dose:

5/sex/group (low and mid doses); 10/sex/group (control, mid high and high doses).

Control animals:

yes, plain diet

Details on study design:

- Dose selection rationale: The concentrations of Ruthenium chloride in diet were selected based on the available data and information from a preliminary dose range finding study [CiToxLAB study code 15/266-100PE].
- Rationale for animal assignment (if not random): Randomisation based on body weights.
- Post-exposure recovery period in satellite groups: 5/sex/group, 28-day daily treatment with control, mid high or high dose. 14-day recovery period.
- Section schedule rationale (if not random): Not specified.

Positive control:

None.

Examinations

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS: Yes.
- Time schedule: At least once a day, at approximately the same time. Animals were inspected for signs of morbidity and mortality twice daily (at the beginning and end of each day).
- Cage side observations: Not specified.

DETAILED CLINICAL OBSERVATIONS: Yes.
- The animals were monitored for any clinical signs, including pertinent behavioural changes, signs of toxicity including mortality, changes in skin, fur, eyes, mucous membranes, occurrence of secretions and excretions, and autonomic activity (e.g. lachrymation, piloerection, pupil size, unusual respiratory pattern), changes in gait, posture and response to handling as well as the presence of clonic or tonic movements, stereotypies (e.g. excessive grooming, repetitive circling), bizarre behaviour (e.g. selfmutilation, walking backwards), observation of tremors, convulsions, salivation,
diarrhoea, lethargy, sleep or coma.

BODY WEIGHT: Yes.
- Time schedule for examinations: 7 days prior to treatment start (Day -7), then daily until the first day of treatment (Day 0). Twice weekly thereafter, including before necropsy.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study):
- Food consumption for each animal determined and mean daily diet consumption calculated as g food//day: Yes (by measuring non-consumed diet on a regular basis throughout the study).
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: Yes.

FOOD EFFICIENCY:
- Mean food utilisation (food conversion efficiency) was calculated: weekly body weight gain (g)/weekly food consumption (g).

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): Not specified.

OPHTHALMOSCOPIC EXAMINATION: No.

HAEMATOLOGY: Yes.
- Time schedule for collection of blood: Post-fasting and immediately prior to the scheduled necropsy (Day 28).
- Anaesthetic used for blood collection: Yes (pentobarbital).
- Animals fasted: Yes.
- How many animals: All animals.
- Parameters checked in table 1 were examined.

CLINICAL CHEMISTRY: Yes.
- Time schedule for collection of blood: Post-fasting and immediately prior to the scheduled necropsy (Day 28).
- Animals fasted: Yes.
- How many animals: All animals.
- Parameters checked in table 2 were examined.

URINALYSIS: No.

NEUROBEHAVIOURAL EXAMINATION: Yes.
- Time schedule for examinations: Day 21/22 of test.
- Dose groups that were examined: All animals.
- Battery of functions tested: sensory activity / grip strength.

IMMUNOLOGY: No.

Sacrifice and pathology:

GROSS PATHOLOGY: Yes (all tissues and organs).

HISTOPATHOLOGY: Yes (see table 3). Tissues and organs in Table 3 were retained from all animals. Full histopathology was performed for control and high dose animals, as well as any organs or tissues from other animals with macroscopic abnormalities.

Statistics:

The mean and standard deviations values, for the frequency of treatment-related clinical observations and macroscopic findings were calculated.

Results and discussion

Results of examinations

Clinical signs:

effects observed, non-treatment-related

Description (incidence and severity):

Dark faeces was observed in the mid high (5000 ppm) and high dose (15,000 ppm) animals from Day 11 to Day 30. This was considered to be a non-adverse observation. No signs of toxicity were observed.

Mortality:

no mortality observed

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

At completion of the feeding period (Day 27), the mean body weight gain of high dose (15,000 ppm) males was about 23% lower than controls. After a 14-day recovery period, these changes were completely recovered. No dose-related or statistically significant changes in body weight gain were observed in the other treated males, or any treated females. The mean body weight and body weight gain of females was more variable than for the males, but there was no clear evidence of a treatment-related effect.

Food consumption and compound intake (if feeding study):

no effects observed

Food efficiency:

effects observed, treatment-related

Description (incidence and severity):

Statistically significantly lower (about 24%) food utilisation was observed in high dose (15,000 ppm) males dues to reduced body weight gain. No effects in other treated males or any treated females.

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

not examined

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

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

no effects observed

Gross pathological findings:

no effects observed

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

no effects observed

Histopathological findings: neoplastic:

no effects observed

Other effects:

not specified

Effect levels

Key result

Dose descriptor:

NOAEL

Effect level:

5 000 ppm

Based on:

test mat.

Sex:

male/female

Basis for effect level:

body weight and weight gain

Target system / organ toxicity

Critical effects observed:

not specified

Applicant's summary and conclusion

Conclusions:

In an OECD 407-compliant 28-day repeated-dose oral toxicity study, rats were fed diets containing 0, 500, 1500, 5000 or 15,000 ppm ruthenium trichloride hydrate (providing up to approximately 1238 mg/kg bw/day). Males in the highest dose group had reduced body weight gain and reduced food conversion efficiency compared to controls. There were no other adverse effects in these animals, and no effects on clinical signs, pathology, organ weights or histopathology in any treated animals. Consequently, the NOAEL is 5000 ppm (providing approximately 407 mg/kg bw/day).

Executive summary:

In an OECD 407 repeated-dose oral toxicity study, conducted according to GLP, groups of Wistar rats were fed diets containing ruthenium trichloride hydrate for 28 days. Diets were formulated to provide 0, 500, 1500, 5000 or 15,000 ppm of the test material (equivalent to approximately 0, 40.6, 124, 407 or 1238 mg/kg bw/day). Male rats recieving the highest dietary concentration (15,000 ppm, approximately 1238 mg/kg bw/day) showed reduced body weight gain compared to controls. These animals also demonstrated a statistically significant reduction in food conversion efficiency. No other significant differences in body weight or body weight gain were reported, although high dose females showed lower body weight gain for the first week only.

There were no adverse effects on food consumption, clinical signs of toxicity, behaviour (functional neurological assessment), haematology, clinical biochemistry, gross pathology, organ weights or histopathology.

The no-observed-adverse-effect level (NOAEL) of ruthenium trichloride hydrate in rats following dietary exposure was 5000 ppm (approximately 407 mg/kg bw/day), on the basis of reduced body weight gain in treated males.