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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
Study period:
2012
Reliability:
1 (reliable without restriction)

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2012
Report Date:
2012

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Type of assay:
bacterial reverse mutation assay

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
other: aqueous solution
Details on test material:
- Name of test material (as cited in study report): IBP1-Na
- Stability under test conditions: Stable
- Storage condition of test material: ambient

Method

Target gene:
Strain Designation Histidine Gene Locus Affected Additional Mutations Type of Mutation Detected
Repair LPS Plasmids
TA 98#1 his D 3052 uvr B- rfa- pKM 101 Frameshift
TA 100#1 his G 46 uvr B- rfa- pKM 101 Base-pair substitution
TA 102#1 his G 428 wild-type rfa- pKM 101 / pAQ1 Base-pair substitution
TA 1535#2 his G 46 uvr B- rfa- none Base-pair substitution
TA 1537#2 his C 3076 uvr B- rfa- none Frameshift


rfa-: partial loss of lipopolysaccharide (LPS) barrier that causes increased permeability to macromolecules
uvr B-: loss of DNA excision repair system
pKM 101: R-factor plasmid, thought to cause an increased error-prone DNA repair
pAQ1: plasmid, carrier of tetracycline resistance
#1 resistance to Ampicillin
#2 non-resistance to Ampicillin
Species / strain
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and TA 102
Metabolic activation:
with and without
Metabolic activation system:
a microsomal preparation derived from Aroclor 1254-induced rat liver.
Test concentrations with justification for top dose:
31.6, 100, 316, 1000, 3160 and 5000 µg/plate
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: aqua ad iniectabilia ( Batch no. 123618002; B. Braun Melsungen AG, 34212 Melsungen, Germany)
Controlsopen allclose all
Untreated negative controls:
yes
Remarks:
aqua ad iniectabilia
Positive controls:
yes
Positive control substance:
sodium azide
Remarks:
in aqua ad iniectabilia (10 µg/plate) : TA 1535, TA 100
Untreated negative controls:
yes
Remarks:
aqua ad iniectabilia
Positive controls:
yes
Positive control substance:
2-nitrofluorene
Remarks:
in DMSO (10 µg/plate) : TA 98
Untreated negative controls:
yes
Remarks:
aqua ad iniectabilia
Positive controls:
yes
Positive control substance:
9-aminoacridine
Remarks:
in ethanol, abs. (100 µg/plate) : TA 1537
Untreated negative controls:
yes
Remarks:
aqua ad iniectabilia
Positive controls:
yes
Positive control substance:
mitomycin C
Remarks:
in DMSO (10 µg/plate) : TA 102
Untreated negative controls:
yes
Remarks:
aqua ad iniectabilia
Positive controls:
yes
Positive control substance:
benzo(a)pyrene
Remarks:
in DMSO (10 µg/plate) : TA 98, TA 102, TA 1537
Untreated negative controls:
yes
Remarks:
aqua ad iniectabilia
Positive controls:
yes
Positive control substance:
other: 2-amino-anthracene
Remarks:
in DMSO (2 µg/plate) : TA 100, TA 1535
Details on test system and experimental conditions:
METHOD OF APPLICATION: in agar (plate incorporation)

DURATION
- Preincubation period: 20 minutes at 37°C
- Exposure duration: 48-72 hours at 37°C
Evaluation criteria:
a test item is considered to show a positive response if
- the number of revertants is significantly increased (p  0.05, U-test according to MANN and WHITNEY, see COLQUHOUN, D.: Lectures on Biostatistics, Clarendon Press, Oxford (1971)) compared with the solvent control to at least 2-fold of the solvent control for TA 98, TA 100 and TA 102 and 3-fold of the solvent control for TA 1535 and TA 1537 in both independent experiments;
- in addition, a significant (p  0.05) concentration (log value)-related effect (Spearman’s rank correlation coefficient, see COLQUHOUN, D.: Lectures on Biostatistics, Clarendon Press, Oxford (1971)) is observed;
- positive results have to be reproducible and the histidine independence of the revertants has to be confirmed by streaking random samples on histidine-free agar plates.

Results and discussion

Test results
Species / strain:
S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and TA 102
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information):
negative with metabolic activation
negative without metabolic activation

Under the present test conditions IBP1-Na, tested up to a concentration of 5000 µg i-Butyl-dtp-Na/plate, caused no mutagenic effect in the Salmonella typhimurium strains TA 98, TA 100, TA 102, TA 1535 and TA 1537 neither in the plate incorporation test nor in the preincubation test each carried out without and with metabolic activation.
Remark: the test solution contains NaOH due to the production method.