Ruthenium trichloride hydrate

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in bacteria

Remarks:

Type of genotoxicity: gene mutation

Type of information:

experimental study

Adequacy of study:

key study

Study period:

16 Sep - 9 Nov 2015

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:

2016

Report date:

2016

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

no

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

Ruthenium trichloride hydrate

EC Number:

604-667-4

Cas Number:

14898-67-0

Molecular formula:

RuCl3.xH2O

IUPAC Name:

Ruthenium trichloride hydrate

Test material form:

solid: particulate/powder

Remarks:

migrated information: powder

Method

Target gene:

histidine

Species / strain

Species / strain / cell type:

S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and TA 102

Additional strain / cell type characteristics:

not applicable

Metabolic activation:

with and without

Metabolic activation system:

mammalian liver post-mitochondrial fraction (S9) prepared from male Sprague Dawley rats induced with Aroclor 1254

Test concentrations with justification for top dose:

Range-finder experiment and mutation experiment 1: 5, 16, 50, 160, 500, 1600 and 5000 µg/plate.
Mutation experiment 2: 160, 300, 625, 1250, 2500 and 5000 µg/plate.

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: purified water
- Justification for choice of solvent/vehicle: well-known solvent/vehicle

Controls

Controls 1

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

2-nitrofluorene

Remarks:

TA98 -S9 5 µg/plate

Controls 2

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

sodium azide

Remarks:

TA100 TA1535 -S9 2 µg/plate

Controls 3

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

9-aminoacridine

Remarks:

TA1537 -S9 50 µg/plate

Controls 4

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

mitomycin C

Remarks:

TA102 -S9 0.2 µg/plate

Controls 5

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

benzo(a)pyrene

Remarks:

TA98 +S9 10 µg/plate

Controls 6

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: 2-aminoanthracene

Remarks:

TA100 TA1535 TA1537 +S9 5 µg/plate; TA102 +S9 20 µg/plate

Details on test system and experimental conditions:

METHOD OF APPLICATION: Toxicity Range-Finder Experiment, Mutation Experiment 1: in agar (plate incorporation); Mutaion Experiment 2: preincubation

DURATION
- Preincubation period: Mutation Experiment 2: 20 minutes
- Exposure duration: 3 days

DETERMINATION OF CYTOTOXICITY
- Method: plates were examined for evidence of toxicity to the background lawn

Evaluation criteria:

For valid data, the test article was considered to be mutagenic if:
1. A concentration related increase in revertant numbers was ≥1.5-fold (in strain TA102), ≥2-fold (in strains TA98 and TA100) or ≥3-fold (in strains TA1535 and TA1537) the concurrent vehicle control values
2. The positive trend/effects described above were reproducible.
The test article was considered positive in this assay if both of the above criteria were met.
The test article was considered negative in this assay if [n]either of the above criteria were met.

Statistics:

None

Results and discussion

Test results

Test results 1

Species / strain:

S. typhimurium TA 1535, TA 1537, TA 98 and TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

In Mutation Experiment 1: from 500 to 5000 µg/plate -S9 and from 1600-5000 µg/plate +S9; Mutation Experiment 2: 2000 and/or 5000 µg/plate -S9 and +S9

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

Test results 2

Species / strain:

S. typhimurium TA 102

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

In Mutation Experiment 1: from 500 to 5000 µg/plate -S9 and from 1600-5000 µg/plate +S9; Mutation Experiment 2: 2000 µg/plate -S9 and +S9

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
- Effects of pH: pH of 50 mg/mL stock formulations was 0.79-0.93.
- Effects of osmolality: no data
- Water solubility: Preliminary solubility data indicated that Ruthenium (III) chloride hydrate was soluble in water for irrigation (purified water) at concentrations equivalent to at least 50 mg/mL.
- Precipitation: Toxicity Range-Finder Experiment and Mutation Experiment 1: no precipitation; Mutation Experiment 2: precipitation at 625 µg/plate and above in all strains in the presence of S9 only

RANGE-FINDING/SCREENING STUDIES: An initial toxicity Range-Finder Experiment was carried out in the absence and in presence of S9 in strains TA98, TA100 and TA102, using final concentrations of Ruthenium (III) chloride hydrate at 5, 16, 50, 160, 500, 1600, and 5000 μg/plate, plus vehicle and positive controls. Following these treatments evidence of toxicity was observed at 1600 and/or 5000 μg/plate in all strains in the absence and presence of S9.

COMPARISON WITH HISTORICAL CONTROL DATA: Mean vehicle control counts fell within the laboratory’s historical ranges; mean positive control counts fell within or slightly above the laboratory's historical control ranges, except for Mutation Experiment 2, TA1535 +S9 for which the mean positive control count was 115.7 and the historical control range was 127-398 (the effect was, nevertheless, clearly positive).

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'.

Applicant's summary and conclusion

Conclusions:

In a guideline study, to GLP, ruthenium (III) chloride hydrate did not induce mutations in five histidine-requiring strains of Salmonella typhimurium, when tested up to 5 mg/plate in the absence or presence of a rat liver metabolic activation system (S9).

Executive summary:

Ruthenium trichloride hydrate was tested in a bacterial reverse mutation (Ames) assay, conducted according to OECD Test Guideline 471 and to GLP.

The test substance was assayed in five histidine-requiring strains (TA98, TA100, TA1535, TA1537 and TA102) of S. typhimurium, both in the absence and in the presence of metabolic activation (S9), in two separate experiments (each in triplicate). The highest concentrations of test article analysed were up to 5000 μg/plate or up to the limit of cytotoxicity and were determined following a preliminary toxicity range-finder experiment. In experiment 1 a plate incorporation protocol was used; the experiment was repeated using a pre-incubation step. Appropriate vehicle and positive control cultures were included in the test system under each treatment condition.

There was no evidence of mutagenicity in any strain with or without S9 in either experiment. Some evidence of toxicity was observed at 1600 and/or 5000 µg/plate with the plate incorporation protocol and at 1250 and/or 2500 µg/plate with the pre-incubation protocol. Vehicle and positive controls performed as expected. It is concluded that ruthenium trichloride hydrate did not induce mutation in five histidine-requiring strains (TA98, TA100, TA1535, TA1537 and TA102) of S. typhimurium when tested at concentrations up to 5000 μg/plate or up to the limit of toxicity, in the absence and in the presence of S9.