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Please be aware that this old REACH registration data factsheet is no longer maintained; it remains frozen as of 19th May 2023.

The new ECHA CHEM database has been released by ECHA, and it now contains all REACH registration data. There are more details on the transition of ECHA's published data to ECHA CHEM here.

Diss Factsheets

Toxicological information

Genetic toxicity: in vitro

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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
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Already evaluated by the Competent Authorities for Biocides and Existing Substance Regulations.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1994

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Deviations:
no
Principles of method if other than guideline:
The method also addressed the following guidelines:
EC Directive 2000/32 Annex V Test B14
UKEMS Guidelines
GLP compliance:
yes
Type of assay:
bacterial reverse mutation assay

Test material

Constituent 1
Reference substance name:
7758-99-8
Cas Number:
7758-99-8
IUPAC Name:
7758-99-8
Constituent 2
Reference substance name:
Cu2+ as Copper sulphate pentahydrate
IUPAC Name:
Cu2+ as Copper sulphate pentahydrate
Details on test material:
Lot/batch number: A668269 350
Description: blue crystalline solid
Purity: 99.0 - 100.5%
Stability: Stable at room temperature

Method

Target gene:
See evaluation criteria.
Species / strain
Species / strain / cell type:
other: Salmonella typhimurium Strains TA98, TA100, TA1535, TA1537, TA102
Metabolic activation:
with and without
Metabolic activation system:
Aroclor 1254-induced rat liver (Sprague-Dawley male rat) post-mitochondrial fraction (S-9 mix)
Test concentrations with justification for top dose:
1.6, 8, 40, 200, 1000 µg/plate and 50, 100, 200, 400, 800 µg/plate in mutation experiments 1 and 2, respectively.
Vehicle / solvent:
None.
Details on test system and experimental conditions:
Study type: Ames test.

Positive control: Details of the positive controls are in Table 1.

Negative control: Yes, tests carried out with purified water in quintuplicate both with and without metabolic activation.

Administration/Exposure; Application of test substance:
Concentrations: Following a range finding study, two experiments were carried out with concentrations of 1.6, 8, 40, 200 and 1000 µg/l in experiment one and 50, 100, 200, 400 and 800 µg/l in experiment two. The tests were carried out in triplicate.

Way of application: The test article was dissolved in sterile purified water.

Pre-incubation time: Only Experiment two included a pre-incubation step for the tests with metabolic activation. The test substance (or control substance), bacteria and S-9 mix were mixed together and incubated for 1 hour at 37 °C before the addition of 2.5 ml molten agar at 46 °C. Plating of these treatments then proceeded as for normal plate-incorporation procedure.

Examinations:
Number of cells evaluated: Colonies were counted electronically using a Seescan Colony Counter and the background lawn inspected for signs of toxicity.
Evaluation criteria:
With the exception of strain TA102, these strains require biotin as well as histidine for growth. In strain TA102 the critical mutation in the histidine gene is located on a multicopy plasmid pAQ1. This strain is particularly sensitive to the activities of oxidative and cross-linking mutagens. The pKM101 plasmid derivatives (TA98, TA100 and TA102) have increased sensitivity to certain mutagens as the pKM101 codes for an error-prone DNA repair system.
Statistics:
The m-statistic was calculated to check that all the data were Poisson-distributed, and the Dunnett's test was used to compare the counts of each dose with the control. The presence or otherwise of a dose response was checked by linear regression analysis.

Results and discussion

Test results
Species / strain:
other: Salmonella typhimurium Strains TA98, TA100, TA1535, TA1537, TA102
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
cytotoxicity
Remarks:
See additional information on results.
Untreated negative controls validity:
valid
Positive controls validity:
valid
Additional information on results:
Evidence of genotoxicity: There was no evidence of genotoxicity observed in either Experiment 1 or Experiment 2 in the presence or absence of metabolic activation.

Cytotoxicity: Evidence of toxicity was observed in all Experiment 1 treatments of 1000 µg/plate. Some evidence of toxicity was also observed following strain TA102 treatments of 200 µg/plate in the presence of S-9 only. In Experiment 2, toxicity was observed following all treatments (with and without metabolic activation) of 800 µg/plate. Some treatments in the presence of S-9 at lower doses also produced evidence of toxicity. The higher degree of toxicity observed with Experiment 2 treatments of S-9 was attributed to the use of a pre-incubation step, which allowed an enhanced exposure of the bacteria to the test article.
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

It was concluded that copper II sulphate pentahydrate was unable to induce mutation in 5 strains of S. typhimurium, when tested at concentrations extending into the toxic range, in the absence and presence of a rat liver metabolic activation system (S-9 mix).
Executive summary:

Materials and Methods

Copper II sulphate pentahydrate was assayed for mutation in 5-histaidine requiring strains (TA98, TA100, TA1537 and TA102) of Salmonella typhimurium, both in the presence and absence of metabolic activation by Aroclor 1254 -induced rat liver post-mitochondrial fraction (S-9) in 2 separate experiments.  Following a range finding study, two experiments were carried out with concentrations of 1.6, 8, 40, 200 and 1000 µg/l in experiment one and 50, 100, 200, 400 and 800 µg/l in experiment two.  The tests were carried out in triplicate.  Both positive and negative controls were included.

The study complied with the following guidelines and was conducted in accordance with GLP:

OECD Guidelines 471
EC Directive 2000/32 Annex V Test B14
UKEMS Guidelines

 

Results and discussion

None of the dose concentrations in any of the test strains in either the absence or presence of S-9 resulted in an increase in revertant numbers that were statistically significant at the 1% level when analysed using a Dunnett’s test.  It was therefore concluded that copper II sulphate pentahydrate was unable to induce mutation in 5 strains ofS. typhimurium, when tested at concentrations extending to the toxic range, in both the absence and presence of rat liver metabolic activation system.