Caesium chloride

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:

2011-10-11 to 2011-10-28

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP and Guideline compliant study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2011

Report date:

2011

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Version / remarks:

adopted 1997

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)

Version / remarks:

adopted 2008

Deviations:

no

Test guideline 3

Qualifier:

according to guideline

Guideline:

EPA OTS 798.5100 (Escherichia coli WP2 and WP2 UVRA Reverse Mutation Test)

Version / remarks:

August 1998

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

Caesium chloride

EC Number:

231-600-2

EC Name:

Caesium chloride

Cas Number:

7647-17-8

Molecular formula:

ClCs

IUPAC Name:

caesium chloride

Test material form:

solid: crystalline

Method

Target gene:

The Salmonella typhimurium histidine (his) reversion system measures his- to his+ reversions. The Salmonella typhimurium strains are constructed to differentiate between base pair (TA 1535, TA 100) and frameshift (TA 1537, TA 98) mutations. The Escherichia coli WP2 uvrA (trp) reversion system measures trp– to trp+ reversions. The Escherichia coli WP2 uvrA detect mutagens that cause other base-pair substitutions (AT to GC).

Species / strain

Species / strain / cell type:

S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and E. coli WP2

Metabolic activation:

with and without

Metabolic activation system:

Phenobarbital (PB) and ß-naphthoflavone (BNF) induced rat liver S 9-mix

Test concentrations with justification for top dose:

5000; 1581; 500; 158; 50 and 15.8 μg/plate.

Vehicle / solvent:

- Vehicle/solvent used: distilled water
- Justification for choice of solvent/vehicle: This vehicle was compatible with the survival of the bacteria and the S9 activity and was chosen based on the results of the preliminary Solubility Test.

Controls

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

Remarks:

1. DMSO; 2. ultrapure water

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: Without metabolic activation: 4-nitro-1,2-phenylene-diamine (TA98); Sodium azide (TA100); 9-aminoacridine (TA1535); Methyl-methanesulfonate (WP2). With metabolic activation: 2-aminoanthracene (all strains)

Details on test system and experimental conditions:

Standard plate incorporation procedure was performed, as an initial mutation test. Bacteria (cultured in Nutrient broth No.2) were exposed to the test item, both in the presence and absence of an appropriate metabolic activation system.
Molten top agar was prepared and kept at 45 °C. 2 mL of top agar was aliquoted into individual test tubes (3 tubes per controls or concentration levels). The equivalent number of minimal glucose agar plates was properly labelled. The test item and other components were prepared fresh and added to the overlay (45 °C).
This solution was mixed and poured on the surface of minimal agar plates. For activation studies, instead of phosphate buffer, 0.5 mL of the S9 Mix was added to each overlay tube. The entire test consisted of non-activated and activated test conditions (without S9 Mix and with addition of S9 Mix) and each of them with the addition of negative and positive controls.
For the pre-incubation method, before overlaying the test item, the bacterial culture and the S9 Mix or phosphate buffer was added into the appropriate tubes, providing direct contact between bacteria and the test item (in its solvent). These tubes were gently mixed and incubated for 20 min at 37 ºC using a shaker. After the incubation the content of the tubes was added to the molten top agar prior to pouring onto the surface of minimal agar plates.
After solidification the plates were inverted and incubated at 37 °C for at least 48 hours in the dark.
The colony numbers on the control, positive control and the test plates were determined, the mean values, standard deviations and the mutation rates were calculated.

Evaluation criteria:

A test item is considered mutagenic if:
- a dose-related increase in the number of revertants occurs and/or;
- a reproducible biologically relevant positive response for at least one of the dose groups occurs in at least one strain with or without metabolic activation.
An increase is considered biologically relevant if:
- in strain TA100 the number of reversions is at least twice as high as the reversion rate of the vehicle control
- in strain TA98, TA1535, TA1537 and Escherichia coli WP2 uvrA the number of reversions is at least three times higher than the reversion rate of the vehicle control.

Statistics:

not applicable

Results and discussion

Test results

Species / strain:

S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and E. coli WP2

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity nor precipitates, but tested up to recommended limit concentrations

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'.

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information):
negative

The test item is considered non-mutagenic in the bacterial reverse mutation assay.

Executive summary:

In compliance with the OECD Guideline No. 471 and EU Method B.13/14, five bacterial strains, Salmonella typhimurium TA 98, TA 100, TA 1535, TA 1537 and Escherichia coli WP2 uvrA were used to investigate the mutagenic potential of cesium chloride. Following concentrations were tested in two independent experiments composed of an initial mutation test (plate incorporation test) and a conformation mutation test (pre-incubation test): 5000, 1581, 500, 158, 50 and 15.8 μg test item/plate. Each assay was conducted with and without metabolic activation (S9 Mix). The concentrations, including the respective positive and negative controls, were tested in triplicate.

No substantial increases or decreases were observed in revertant colony numbers of any of the five test strains following treatment with cesium chloride at any concentration level, either in the presence or absence of metabolic activation (S9 Mix) in the performed experiments. Sporadic increases in revertant colony numbers compared to the vehicle control values and/or revertant colony numbers above the actual historical control data ranges were observed in both independently performed main experiments. However, there was no tendency of higher mutation rates with increasing concentrations beyond the generally acknowledged border of biological relevance in the performed experiments.

Thus, it can be concluded that the test item did not induce gene mutations by frameshift or base-pair substitution in the genome of the strains used. Therefore, cesium chloride is considered non-mutagenic in this bacterial reverse mutation assay.