Cyclohexanol, 4-[[(2-amino-3,5-dibromophenyl)methyl]amino]-, hydrobromide (1:1), trans-

Genetic toxicity: in vitro

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:

1999-01-22 to 1999-01-25

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: The study was conducted under GLP conditions in accordance with the official OECD guideline No 471 (1997).

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1999

Report date:

1999

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Version / remarks:

(adopted July 1997)

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)

Version / remarks:

(adopted December 1992)

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

Mucosolvan HBr

IUPAC Name:

Mucosolvan HBr

Details on test material:

- Name of test material (as cited in study report): Mucosolvan HBr
- Physical state: Beige solid
- Stability in vehicle: Dimethylsulfoxide: not indicated

Method

Target gene:

Strain: TA98; Histidin mutation: hisD3052/R-factor; Mutation type: frameshift
Strain: TA100; Histidin mutation: hisG46/R-factor; Mutation type: base-pair substitutions

Both tester strains contained the following additional mutations:
rfa: deep rough (defective lipopolysaccharide cellcoat)
gal: mutation in the galactose metabolism
chl: mutation in nitrate reductase
bio: defective biotin synthesis
uvrB: loss of the excision repair system (deletion of the ultraviolet-repair B gene)

Species / strain

Species / strain / cell type:

S. typhimurium, other: TA 100 and TA 98

Details on mammalian cell type (if applicable):

- Properly maintained: yes

Metabolic activation:

with and without

Metabolic activation system:

S9

Test concentrations with justification for top dose:

3, 10, 33, 100, 333, 1000, 3330, 5000 µg/plate

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: DMSO

Controls

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

methylmethanesulfonate

other: daunomycine, 2-aminoanthracene

Details on test system and experimental conditions:

METHOD OF APPLICATION: in agar (plate incorporation)

DURATION
- Exposure duration: 48 h

NUMBER OF REPLICATIONS: 3

DETERMINATION OF CYTOTOXICITY
- Method: background growth, number of revertants

Evaluation criteria:

A test substance is considered negative (not mutagenic) in the test if: The total number of revertants in any tester strain at any concentration is not greater than two times the solvent control value, with or without metabolic activation.
A test substance is considered positive (mutagenic) in the test if: It induces at least a 2-fold, dose related increase in the number of revertants with respect to the number induced by the solvent control in any of the tester strains, either with or without metabolic activation. However, any mean plate count of less than 20 is considered to be not significant.

Results and discussion

Test results

Species / strain:

S. typhimurium, other: TA 100 and TA 98

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'.

Any other information on results incl. tables

The bacterial background lawn was not reduced at all concentrations tested in both tester strains. In tester strain TA 98, an extreme reduction in the number of revertants was observed at the test substance concentration of 5000 µg/plate in the absence of S9 -mix. In the presence of S9 -mix, no decrease in the number of revertants was observed. In tester strain TA 100, an extreme reduction in the number of revertants was observed at test substance concentrations of 3330 and 5000 µg/plate in the absence of S9-mix and at the test substance concentration of 5000 µg/plate in the presence of S9-mx.

Mucosolvan HBR did not precipate in the top agar. Precipitation of the substance was not observed on the plates at the start or at the end of the incubation period in both tester strains.

Mucosolvan HBR did not induce a dose-related increase in the number of revertant colonies both in the absence and the presence of S9-mix.

Table 1: Mutagenic response of Mucosolvan HBR in the Salmonella Typhimurium reverse mutation assay

Dose (µg/plate)	Mean number of revertant (His+) colonies/ 3 replicates plates (+-S.D.) with two strains of Salmonella typhimurium
	TA98		TA100
	Without S9 mix
Positive control	594	+-74	893	+-41
Solvent control	28	+-3	96	+-16
3	32	+-5	82	+-5
10	28	+-8	92	+-6
33	26	+-2	88	+-9
100	30	+-2	91	+-5
333	29	+-2	88	+-19
1000	24	+-4	93	+-9
3330	22	+-2	30	+-6
5000	9	+-3	9	+-7
	With S9 mix
Positive control	738	+-100	2347	+-32
Solvent control	43	+-5	95	+-5
3	35	+-5	97	+-13
10	31	+-7	104	+-12
33	31	+-6	109	+-10
100	35	+-6	106	+-23
333	47	+-7	104	+-3
1000	48	+-9	103	+-11
3330	43	+-4	77	+-9
5000	31	+-8	21	+-7

Solvent control; 0.1 ml dimethylsulphoxide

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information):
negative

Based on the results of this study it is concluded that Mucosolvan HBr is not mutagenic in the Salmonella typhimurium reverse mutation assay.