Reaction mass of barium chromate, copper dichromium tetraoxide and copper oxide

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in mammalian cells

Remarks:

Type of genotoxicity: gene mutation

Type of information:

experimental study

Adequacy of study:

key study

Study period:

not indicated

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Publication with unknown GLP study status. Non-guideline assay with several limitations with regard to sensitivity. The sensitivity of the test is however validated by the positive result obtained. Very limited details on test item.

Data source

Reference

Reference Type:

publication

Title:

Chromate-Induced Epimutations in Mammalian Cells

Author:

Klein CB, Su L, Bowser D et al

Year:

2002

Bibliographic source:

Environ Health Perspect 110(suppl 5) :739-743 (2002)

Report date:

2002

Materials and methods

Test guideline

Qualifier:

no guideline followed

Principles of method if other than guideline:

These transgenic cells allow the characterization of a mixed spectrum of genotoxic outcomes that may include genetic mutations, transgene deletions, and epigenetic gene silencing caused by aberrant DNA hypermethylation.

GLP compliance:

not specified

Type of assay:

mammalian cell gene mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

Barium chromate

EC Number:

233-660-5

EC Name:

Barium chromate

Cas Number:

10294-40-3

Molecular formula:

Ba.CrH2O4

IUPAC Name:

Barium chromate

Details on test material:

- Name of test material (as cited in study report): Barium chromate
- Molecular formula (if other than submission substance): BaCrO4
- Physical state: solid particles
- Granulometry: text states "sized at 0.4–0.6 μM", the intended meaning was probably "size 0.4–0.6 μm" ?

- No other details available

Method

Target gene:

GPT = guanine-hypoxanthine phosphoribosyltransferase

Species / strain

Species / strain / cell type:

other: Chinese hamster G12 lung cells

Details on mammalian cell type (if applicable):

G12 cell line is V79-derived and contains a bacterial GPT reporter gene in its DNA: transgenic cell line

Metabolic activation:

without

Test concentrations with justification for top dose:

0.05–0.25 μg/cm2 (lower concentrations seem to have been tested for cytotoxicity)

Vehicle / solvent:

complete F12 medium

Controls

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

no

Remarks:

(but test validated by positive result with test item)

Positive control substance:

no

Details on test system and experimental conditions:

METHOD OF APPLICATION: suspension in medium

DURATION
- Exposure duration: 24 hr at 37°C in complete F12 medium to allow dissolution of Cr and/or phagocytic uptake by cells
- Expression time (cells in growth medium): 7 days in nonselective F12 medium
- Selection time (if incubation with a selection agent): not indicated
- Fixation time (start of exposure up to fixation or harvest of cells): not indicated

SELECTION AGENT (mutation assays): 10 μg/mL 6TGF12 medium (selects the 6-thioguanine resistance)

NUMBER OF REPLICATIONS: 2 or 3 per dose

NUMBER OF CELLS EVALUATED: not indicated

DETERMINATION OF CYTOTOXICITY
- Method: Clonal cell survival

OTHER EXAMINATIONS:
After the classical assay, 6TG resistant mutants/epimutants were further evaluated for the nature of the genotoxic event:
- Deletion of the transgene was investigated by extraction of high MW DNA from 0.5-1 × 10^7 resistant cells and PCR.
- Gene silencing caused by DNA hypermethylation was investigated by Southern blot for the transgene and its promoter region.

Evaluation criteria:

Increase in mutant frequency over controls

Statistics:

not performed

Results and discussion

Test results

Species / strain:

other: Chinese hamster G12 lung cells

Metabolic activation:

without

Genotoxicity:

positive

Remarks:

at 0.15 and 0.20 µg/cm2

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

-40% survival at 0.20 µg/cm2

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

not applicable

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
RANGE-FINDING/SCREENING STUDIES:
COMPARISON WITH HISTORICAL CONTROL DATA:
all no data.

CYTOTOXICITY:
Dose-related decrease in survival; >80% up to 0.10 µg/cm2, 75% at 0.15 µg/cm2, 60% at 0.20 µg/cm2 (as assessed from graph)

MUTATION:
BaCrO4 was mutagenic in G12 cells, with a maximal mutation peak (3.5× background; 3.2x vehicle control as assessed from graph) at 0.15 μg/cm2 and a decline at next dose (2.1x vehicle control as assessed from graph).

FURTHER EXPLANATORY ASSAYS:
- 33% (8/24) of mutant G12 cells showed transgene deletion at PCR
- BaCrO4 did not induce any DNA methylation changes in G12 cells

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):
positive without metabolic activation no metabolic activation tested

Barium Chromate induced gene mutation (and cytotoxicity) in transgenic Chinese Hamster lung cells. The mutagenicity could not be explained by the cytotoxicity.

Executive summary:

When tested in Chinese Hamster lung G12 trangenic cells, Barium Chromate induced concentration-dependent increases in cytotoxicity and gene mutation. The mutation was mainly due to deletion (33% of mutants). The results were characteristic of the mutagenesis profiles in various mammalian cells by Cr.