Potassium iodide

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro cytogenicity / micronucleus study

Remarks:

Type of genotoxicity: chromosome aberration

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

data from handbook or collection of data

Justification for type of information:

Data is from peer reviewed publication

Data source

Reference

Reference Type:

publication

Title:

Gene mutation toxicity study of the test chemical

Author:

Poul et al

Year:

2004

Bibliographic source:

Food and Chemical Toxicology

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

other: Refer below principle

Principles of method if other than guideline:

Genotoxic effects of the test chemical was evaluated in vitro using the cytokinesis-block micronucleus assay on CHO cells

GLP compliance:

not specified

Type of assay:

in vitro mammalian cell micronucleus test

Test material

Test material information

Constituent 1

Reference substance name:

Potassium iodide

EC Number:

231-659-4

EC Name:

Potassium iodide

Cas Number:

7681-11-0

Molecular formula:

IK

IUPAC Name:

potassium iodide

Details on test material:

- Name of test material: potassium iodide
- Molecular formula : KI
- Molecular weight : 166.0 g/mol
- Substance type: Inorganic
- Physical state: Solid

Method

Target gene:

No data

Species / strain

Species / strain / cell type:

Chinese hamster Ovary (CHO)

Details on mammalian cell type (if applicable):

Details on mammalian cell line
- Type and identity of media: The cell line was grown at 37 C in a humidified atmosphere at
5% CO2 in air, in HAM’S F12 medium with L-glutamine supplemented with 10% fetal calf serum (FCS), penicillin (50 UI/ml) and streptomycine (50 µg/ml). Cells were subcultured 24 h before treatment.
- Properly maintained: Yes
- Periodically checked for Mycoplasma contamination: No data
- Periodically checked for karyotype stability: No data
- Periodically "cleansed" against high spontaneous background: No data

Additional strain / cell type characteristics:

not specified

Cytokinesis block (if used):

No data

Metabolic activation:

not specified

Metabolic activation system:

no data

Test concentrations with justification for top dose:

0.625, 1.25, 2.5, 5 and 10 mM

Vehicle / solvent:

culture medium- Vehicle(s)/solvent(s) used: Culture medium
- Justification for choice of solvent/vehicle: The test chemical was soluble in culture medium

Controls

Untreated negative controls:

yes

Negative solvent / vehicle controls:

not specified

True negative controls:

not specified

Positive controls:

yes

Positive control substance:

methylmethanesulfonate

Remarks:

MMS (30 µg/ml)

Details on test system and experimental conditions:

Cell culture procedure
Chinese hamster ovary (CHO-K1) cells were purchased from Eurobio (France). They were routinely maintained from stocks stored in liquid nitrogen. CHO cells were grown at 37 C in a humidified atmosphere at 5% CO2 in air, in HAM’S F12 medium with l-glutamine supplemented with 10% fetal calf serum (FCS), penicillin (50 UI/ml) and streptomycine (50 mg/ml). Cells were subcultured 24 h before treatment.

Cell treatment
In preliminary cytotoxicity assays, CHO cells were exposed for 1 h to the test compounds at concentrations ranging from 0.001 to 5 mg/ml.
In the alkaline comet assay and in the cytokinesisblock micronucleus assay, cells were exposed for 3 h to concentrations of the test compounds of 0.625, 1.25, 2.5,5 and 10 mM. Tests compounds and MMS (30 mg/ml) were dissolved the culture medium. Etoposide (0.5 mg/ml) was dissolved in DMSO.

METHOD OF APPLICATION: in medium
- Cell density at seeding (if applicable): 150000 cells/well

DURATION
- Preincubation period: No data
- Exposure duration: 3 hrs
- Expression time (cells in growth medium): 20 hrs
- Selection time (if incubation with a selection agent): No data
- Fixation time (start of exposure up to fixation or harvest of cells): No data

SELECTION AGENT (mutation assays): No data

SPINDLE INHIBITOR (cytogenetic assays): No data

STAIN (for cytogenetic assays): Acridine orange

NUMBER OF REPLICATIONS: Duplicate

METHODS OF SLIDE PREPARATION AND STAINING TECHNIQUE USED: Cells were fixed with cold methanol, stained with acridine orange (62.5 mg/ml) for 5 min and mounted in Sorensen buffer. Slides were coded and blindly examined under an epifluorescence microscope at 1000X magnification under oil immersion.

NUMBER OF CELLS EVALUATED: One thousand (1000) binucleated cells were scored
for each slide.

NUMBER OF METAPHASE SPREADS ANALYSED PER DOSE (if in vitro cytogenicity study in mammalian cells): No data

CRITERIA FOR MICRONUCLEUS IDENTIFICATION: MN should be morphologically identical to but smaller than nuclei, their diameter usually varied between 1/6th and 1/3rd of the mean diameter of the main nuclei. MN should be readily distinguished and not be linked to the main nuclei via nucleoplasmic bridges. Cells showing chromatin condensation or nuclear fragmentation with an intact cytoplasmic membrane were classified as apoptotic cells.

DETERMINATION OF CYTOTOXICITY
- Method: mitotic index; cloning efficiency; relative total growth; other: Yes, Cytotoxicity was measured by the Binucleate cell ratio between treated and control slides
- Any supplementary information relevant to cytotoxicity: No data

OTHER EXAMINATIONS:
- Determination of polyploidy: No data
- Determination of endoreplication: No data
- Methods, such as kinetochore antibody binding, to characterize whether micronuclei contain whole or fragmented chromosomes (if applicable): No data

- OTHER: No data

Rationale for test conditions:

No data

Evaluation criteria:

MN should be morphologically identical to but smaller than nuclei, their diameter usually varied between 1/6th and 1/3rd of the mean diameter of the main nuclei. MN should be readily distinguished and not be linked to the main nuclei via nucleoplasmic bridges. Cells showing chromatin condensation or nuclear fragmentation with an intact cytoplasmic membrane were classified as apoptotic cells.

Statistics:

In the cytokinesis-block micronucleus assay, data were expressed as the percentage of binucleated cells with micronuclei. Comparisons between control and treated cell cultures were made using ANOVA and Dunnett’s one sided test.

Results and discussion

Test results

Species / strain:

Chinese hamster Ovary (CHO)

Metabolic activation:

not specified

Genotoxicity:

negative

Remarks:

Potassium iodate did not induce any increase in the frequency of MNBN cells for doses ranging from 0.625 to 10 mM.

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

not specified

Untreated negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
- Effects of pH: No data
- Effects of osmolality: No data
- Evaporation from medium: No data
- Water solubility: No data
- Precipitation: No data
- Definition of acceptable cells for analysis: No data
- Other confounding effects: No data

RANGE-FINDING/SCREENING STUDIES: In preliminary cytotoxicity assays, CHO cells were
exposed for 1 h to the test compounds at concentrations ranging from 0.001 to 5 mg/ml.

CYTOKINESIS BLOCK (if used)
- Distribution of mono-, bi- and multi-nucleated cells: No data

NUMBER OF CELLS WITH MICRONUCLEI
- Number of cells for each treated and control culture: No data
- Indication whether binucleate or mononucleate where appropriate: No data

HISTORICAL CONTROL DATA (with ranges, means and standard deviation and confidence interval (e.g. 95%)
- Positive historical control data: No data
- Negative (solvent/vehicle) historical control data: No data

ADDITIONAL INFORMATION ON CYTOTOXICITY:
- Measurement of cytotoxicity used: No data
- Other observations when applicable: No data

Applicant's summary and conclusion

Conclusions:

The test chemical did not induce any increase in the frequency of MNBN cells for doses ranging from 0.625 to 10 mM in the micronucleus assay in the CHO cell line and hence it is not liekly to classify as a gene mutant in vitro.

Executive summary:

Genotoxic effects of the test chemical was evaluated in vitro using the cytokinesis-block micronucleus assay on CHO cells. The study was performed using CHO cells. The test chemical was dissolved in culture medum and used at dose level of 0.625, 1.25, 2.5, 5 and 10 mM. In preliminary cytotoxicity assays, CHO cells were exposed for 1 h to the test compound at concentrations ranging from 0.001 to 5 mg/ml. The cells were exposed for 3 h to concentrations of the test compounds of 0.625, 1.25, 2.5, 5 and 10 mM. Test compound and MMS (30 mg/ml) were dissolved the culture medium. Exponentially growing CHO-K1 cells were plated in a six-well plate on glass coverslips (1.5 X 10⁵ cells/well) and cultured 24 h prior to compound treatment. Duplicate coverslips were established for each experiment, and at least two independent experiments were performed. The cells were exposed to the chemicals at different concentrations for 3 h in a FCS free medium. At the end of treatment, cells were washed twice with PBS before a 20 h incubation in fresh medium containing 10% of FCS and 3 mg/ml of cytochalasin B. Thereafter, cells were washed twice with PBS and allowed to recover for 1.5 h in 10% FCS fresh medium. Cells were fixed with cold methanol, stained with acridine orange (62.5 mg/ml) for 5 min and mounted in Sorensen buffer. Slides were coded and blindly examined under an epifluorescence microscope at 1000X magnification under oil immersion. Briefly, the cells should be binucleated (BN) with an intact nuclear membrane and should be situated within the same cytoplasmic boundary. MN should be morphologically identical to but smaller than nuclei, their diameter usually varied between 1/6th and 1/3rd of the mean diameter of the main nuclei. MN should be readily distinguished and not be linked to the main nuclei via nucleoplasmic bridges. Cells showing chromatin condensation or nuclear fragmentation with an intact cytoplasmic membrane were classified as apoptotic cells. One thousand (1000) binucleated cells were scored for each slide. The frequencies of BN, of BN with MN (MNBN) and of apoptotic cells (AP) were estimated. MMS (30 mg/ml), a well known alkylating agent was used as positive control. Cytotoxicity was measured by the BN cell ratio between treated and control slides. Based on the observations made, the test chemical did not induce any increase in the frequency of MNBN cells for doses ranging from 0.625 to 10 mM in the micronucleus assay in the CHO cell line and hence it is not liekly to classify as a gene mutant in vitro.