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Toxicological information

Genetic toxicity: in vitro

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Administrative data

Endpoint:
in vitro gene mutation study in bacteria
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
study well documented, meets generally accepted scientific principles, acceptable for assessment
Justification for type of information:
Data is from peer reviewed journal

Data source

Reference
Reference Type:
publication
Title:
Mutagenicity Tests On Anthelmintics: Microsomal activation of Viprynium Embonate to a mutagen
Author:
D.G. Macphee and D.M. Podger
Year:
1977
Bibliographic source:
Mutation Research, 48 (1977) 307-312

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
other: as metioned below
Principles of method if other than guideline:
A slightly modified version of the procedure recommended by Ames et al. was adopted, in that the test samples were simply placed in "wells" cut out of the agar of a plate previously seeded with the appropriate tester strain.
GLP compliance:
not specified
Type of assay:
bacterial reverse mutation assay

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
solid: particulate/powder
Remarks:
migrated information: powder
Details on test material:
- Name of test material (as cited in study report): Piperazine Citrate - Molecular formula (if other than submission substance): C12H30N6•Cl2H16O14 - Molecular weight (if other than submission substance): 642.76 g/mole - Smiles notation (if other than submission substance): C1CNCCN1.C1CNCCN1.C1CNCCN1.C(C(=O)O)C(CC(=O)O)(C(=O)O)O.C(C(=O)O)C(CC(=O)O)(C(=O)O)O - Substance type: Organic - Physical state: Solid-Purity:99.0% (BP)
Specific details on test material used for the study:
- Name of test material (as cited in study report): Piperazine citrate / tripiperazine dicitrate- Molecular formula: C12H30N6•Cl2H16O14 - Molecular weight: 642.76 g/mole - Substance type: Organic - Physical state: Solid

Method

Species / strain
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98 and TA 100
Details on mammalian cell type (if applicable):
not specified
Additional strain / cell type characteristics:
not specified
Metabolic activation:
with and without
Metabolic activation system:
S-9 mix, which contained per ml: 0.3 ml of S-9 fraction 8 µmol MgCl2, 33 µmol KCl, 5 µmol glucose-6-phosphate, 4 µmol NADP, and 100 µmol sodium phosphate
Test concentrations with justification for top dose:
5000 µg/ml or 50,000 µg/ml.
Vehicle / solvent:
DMSO
Controls
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
True negative controls:
not specified
Positive controls:
not specified
Positive control substance:
not specified
Details on test system and experimental conditions:
METHOD OF APPLICATION: in agar (plate incorporation)DURATION- Preincubation period: 2-3 days- Exposure duration: 24 hrs- Expression time (cells in growth medium): not specified- Selection time (if incubation with a selection agent): not specified- Fixation time (start of exposure up to fixation or harvest of cells): not specifiedSELECTION AGENT (mutation assays): SPINDLE INHIBITOR (cytogenetic assays): not specifiedSTAIN (for cytogenetic assays): not specifiedNUMBER OF REPLICATIONS: not specifiedNUMBER OF CELLS EVALUATED: not specifiedDETERMINATION OF CYTOTOXICITY- Method: mitotic index; cloning efficiency; relative total growth; other:OTHER EXAMINATIONS: not specified- Determination of polyploidy:- Determination of endoreplication:- Other:
Rationale for test conditions:
not specified
Evaluation criteria:
not specified
Statistics:
not specified

Results and discussion

Test results
Key result
Species / strain:
S. typhimurium, other: TA98, TA100, TA1535 and TA1538TA98, TA100, TA1535 and TA1538
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
not specified
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
not specified

Applicant's summary and conclusion

Conclusions:
The test substance piperazine citrate gave negative results for mutagenicity with or without S9 metabolic activation system when tested at concentration of 5000 µg/ml or 50,000 µg/ml in Salmonella typhimurium TA98, TA100, TA1535 and TA1538. Thus the test chemical is not likely to classify as a gene mutant in vitro as per the CLP classification criteria.
Executive summary:

The test substance piperazine citrate was tested for mutagenicity in the Salmonella typhimurium test system. The strains used wereTA98, TA100, TA1535 and TA1538.The test concentrations used were 5000µg/ml or 50,000µg/ml.A slightly modified version of the procedure recommended by Ames et al. was adopted, in that the test samples were simply placed in "wells" cut out of the agar of a plate previously seeded with the appropriate tester strain. Addition of a mixture of rat liver microsomal enzymes and appropriate co-factors ("S-9 mix") to one of the two wells on a single plate allowed a possible requirement for metabolic activation to be recognised. Using this procedure, the test chemical was determined to be non-mutagenic in this system. Controls with DMSO alone and S-9 mix alone also gave negative results. Thus the test chemical is not likely to classify as a gene mutant in vitro as per the CLP classification criteria.