Cuprate(4-), [.mu.-[[3,3'-[(1-oxido-1,2-diazenediyl)bis[[2-(hydroxy-.kappa.O)-4,1-phenylene]-2,1-diazenediyl-.kappa.N1]]bis[4-(hydroxy-.kappa.O)-2,7-naphthalenedisulfonato]](8-)]]di-, ammonium sodium

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:

1991-05-29 and 1991-07-30

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: in accordance to OECD Guideline and EU method, GLP-conform

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1991

Report date:

1991

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Version / remarks:

1983

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

1984

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

FAT 11181/K

IUPAC Name:

FAT 11181/K

Test material form:

other: solid

Method

Species / strain

Species / strain / cell type:

other: S. Typhimurium TA 1535, TA 1537, TA 1538, TA 98 and TA 100

Metabolic activation:

with and without

Metabolic activation system:

S9 rat liver microsomal fraction induced by 500 mg/kg bw Aroclor 1254

Test concentrations with justification for top dose:

10.0; 100.0; 333.3; 1000.0; and 5000.0 ug/plate

Vehicle / solvent:

- Vehicle/solvent used: aqua bidest.
- Justification for choice of solvent/vehicle: The solvent was chosen because of its solubility properties and its relative nontoxicity for the bacteria.

Controls

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

Remarks:

aqua dest. and DMSO

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: with S9: 2-aminoanthracene (strain: TA1535, TA1537, TA1538, TA98, T100); without S9: sodium azide (strain: TA1535, TA100), 4-nitro-o-phenylene-diamine (strain: TA1537, TA1538, TA98)

Details on test system and experimental conditions:

METHOD OF APPLICATION: in agar (plate incorporation)

PRECULTURE: From the thawed ampoules of the strains 0.5 mL bacterial suspension was transferred to 250 mL Erlenmeyer flasks containing 20 mL nutrient medium. The bacterial culture was incubated in a shaking water bath for 10 hours at 37° C.

DURATION
- Exposure duration: 48 hours at 37°C in the dark

SELECTION AGENT: 2.0 % Vogel-Bonner-Glucose-Minimal-Agar

NUMBER OF REPLICATIONS: three plates per strain and dose level; 2 independent experiments

DATA RECORDING: The colonies were counted by using the BIOTRAN III counter (BIOTRONIK, D-6000 Frankfurt, F.R.G,). If precipitation of the test article precluded automatic counting the revertant colonies were counted by hand.

Evaluation criteria:

A test article is considered as positive if either a significant dose-related increase in the number of revertants or a significant and reproducible increase for at least one test concentration is induced. A significant response is described as follows:
A test article is considered as mutagenic if in strain TA 100 the number of reversions is at least twice as high and in strains TA 1535, TA 1537, TA 1538, and TA 98 it is at least three times higher as compared to the spontaneous reversion rate.

Results and discussion

Test results

Test results 1

Species / strain:

S. typhimurium, other: TA 1537 and TA 1538

Metabolic activation:

with and without

Genotoxicity:

positive

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 2

Species / strain:

S. typhimurium TA 98

Metabolic activation:

without

Genotoxicity:

positive

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 3

Species / strain:

S. typhimurium TA 98

Metabolic activation:

with

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 4

Species / strain:

S. typhimurium, other: TA 1535 and TA 100

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

Additional information on results:

RANGE-FINDING/SCREENING STUDIES: The plates with the test article showed normal background growth up to 5000.0 µg/plate in strain TA 98 and TA 100, respectively. According to the dose selection criteria, the test article was tested at the following concentrations:
10.0; 100.0; 333.3; 1000.0; and 5000.0 µg/plate

ADDITIONAL INFORMATION ON CYTOTOXICITY: No toxic effects occurred in the test groups with and without metabolic activation in experiment I and II in all strains used.

Remarks on result:

other: strain/cell type: TA 1537 and TA 1538

Remarks:

Migrated from field 'Test system'.

Any other information on results incl. tables

The results of the main study are shown in table 1 (without S9 mix) and table 2 (with S9 mix).

 

Table 1: without S9 mix

Revertants/plate (mean from three plates)

Dose µg/plate I/II	TA 1535		TA 1537		TA1538		TA98		TA100
	I	II	I	II	I	II	I	II	I	II
Neg. contr.	7	15	10	8	12	15	20	23	77	78
Solv. contr	11	14	9	9	11	12	22	22	85	81
10.0	11	12	11	11	13	13	18	28	79	76
100.0	14	18	9	14	23	27	31	32	80	66
333.3	10	13	13	17	44	49	35	44	77	79
1000.0	10	11	26	21	73	83	49	63	90	91
5000.0	11	15	62	67	162	156	123	100	87	94
Pos. contr.
Sodium azide (10 µg/plate)	1129	932							901	714
4-Nitro-o-phenylene-diamine (50 µg/plate)			158	186	1635	1451	1721	1509

 

Table 2: with S9 mix

Revertants/plate (mean from three plates)

Dose µg/plate I/II	TA 1535		TA 1537		TA1538		TA98		TA100
	I	II	I	II	I	II	I	II	I	II
Neg. contr.	12	15	9	13	22	18	27	34	113	93
Solv. contr	11	11	8	9	14	14	26	36	114	80
10.0	10	12	7	10	15	15	27	30	114	84
100.0	11	14	8	12	14	20	26	34	101	84
333.3	10	13	8	8	29	31	27	29	120	76
1000.0	11	13	12	16	29	31	29	32	112	92
5000.0	12	16	26	31	93	83	76	56	125	100
Pos. contr.
2-Amino-anthracene (2.5 µg/plate)	305	288	140	226	998	934	347	1418	475	998