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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:
1988-05-04 to 1988-06-13
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1988
Report Date:
1988

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Version / remarks:
adopted 26 May 1983
Deviations:
no
GLP compliance:
yes
Type of assay:
bacterial reverse mutation assay

Test material

Reference
Name:
Unnamed
Type:
Constituent

Method

Target gene:
histidine locus
Species / strainopen allclose all
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98 and TA 100
Species / strain / cell type:
S. typhimurium TA 1538
Metabolic activation:
with and without
Metabolic activation system:
S9 mix
Test concentrations with justification for top dose:
Experiment 1: 0, 8, 40, 200, 1000 and 5000 µg/plate
Experiment 2: 1000, 2000, 3000, 4000 and 5000 µg/plate
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: water
- Justification for choice of solvent/vehicle: test substance is water soluble
Controlsopen allclose all
Negative solvent / vehicle controls:
yes
Positive controls:
yes
Positive control substance:
9-aminoacridine
2-nitrofluorene
sodium azide
Remarks:
without metabolic activation
Positive controls:
yes
Positive control substance:
other: 2-aminoanthracene
Remarks:
with metabolic activation
Details on test system and experimental conditions:
METHOD OF APPLICATION: in agar (plate incorporation)

DURATION
- Exposure duration: at least 2 days

NUMBER OF REPLICATIONS: triplicate plates were used for testsubstance concentrations,quintuplicate plates for negative controls ,
two independent experiments

DETERMINATION OF CYTOTOXICITY
- Method: background lawn



Evaluation criteria:
Validity criteria:
(i) the mean negative control counts fell within the normal range.
(ii) the positive control chemicals induced clear increases in revertant numbers.
(iii) no more than 5% of the plates were lost through contamination or some other unforseen event.

Evaluation criteria:
A two-fold increase in TA98 or TA100 revertants, and a three-fold increase in TA1535, TA1537 or TA1538 revertants, from concurrent controls, indicates a positive response.
Statistics:
none

Results and discussion

Test resultsopen allclose all
Key result
Species / strain:
S. typhimurium TA 1535
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity nor precipitates, but tested up to recommended limit concentrations
Vehicle controls validity:
valid
Positive controls validity:
valid
Key result
Species / strain:
S. typhimurium TA 1537
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity nor precipitates, but tested up to recommended limit concentrations
Vehicle controls validity:
valid
Positive controls validity:
valid
Key result
Species / strain:
S. typhimurium TA 1538
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity nor precipitates, but tested up to recommended limit concentrations
Vehicle controls validity:
valid
Positive controls validity:
valid
Key result
Species / strain:
S. typhimurium TA 98
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity nor precipitates, but tested up to recommended limit concentrations
Vehicle controls validity:
valid
Positive controls validity:
valid
Key result
Species / strain:
S. typhimurium TA 100
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity nor precipitates, but tested up to recommended limit concentrations
Vehicle controls validity:
valid
Positive controls validity:
valid
Additional information on results:
TEST-SPECIFIC CONFOUNDING FACTORS
- Water solubility: soluble
- Precipitation: no

RANGE-FINDING/SCREENING STUDIES:
0, 8, 40, 200, 1000 and 5000 µg/plate - no toxicity was observed

COMPARISON WITH HISTORICAL CONTROL DATA:
all data within historical control data

No substantial increases in revertant colony numbers of any of the five tester strains were observed following treatment with the test substance at any dose level, either in the presence or absence of metabolic activation (S9 mix). There was also no tendency of higher mutation rates with increasing concentrations in the range below the generally acknowledged border of significance.

Applicant's summary and conclusion

Conclusions:
Guanidine hydrochloride is considered to be negative in the reverse gene mutation assay in bacteria (S. typhimurium strains TA 98, TA 100, TA 1535, TA 1537 and TA 1538), when tested up to the limit concentration of 5000 µg/plate in the presence and absence of mammalian metabolic activation.
Executive summary:

In a reverse gene mutation assay in bacteria according to OECD guideline 471, 1983, strains TA 98, TA 100, TA 1535, TA 1537 and TA 1538 of S. typhimurium were exposed to Guanidine hydrochloride (98.5% a.i.) in water at concentrations of at concentrations of 8, 40, 200, 1000 and 5000 µg/plate in the first experiment and 1000, 2000, 3000, 4000, 5000 µg/plate in the second experiment in the presence and absence of mammalian metabolic activation (rat liver S9-mix) using the plate co-incubation method.

Guanidine hydrochloride was tested up to limit concentrations of 5000 µg/plate. No cytotoxicity was observed and no increase in the number of revertants were observed in all tester strains, with or without metabolic activation. The positive controls induced the appropriate responses in the corresponding strains and activity of metabolising system was confirmed.

There was no evidence of induced mutant colonies over background.

This study is classified as acceptable and satisfies the requirement for Test Guideline OECD 471 (1983) for in vitro mutagenicity (bacterial reverse gene mutation) data.