6,15-dihydroanthrazine-5,9,14,18-tetrone

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

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Guideline study with acceptable restriction - No data on the purity of the tested article. - Neither TA 102 nor E. Coli is included in the panel of cells tested. - No data on GLP

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1986

Report date:

1986

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

yes

Remarks:

(neither TA 102 nor E. coli is included in the panel of cells tested; no statistical analysis, no data on purity of the tested article;)

GLP compliance:

not specified

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

6,15-dihydroanthrazine-5,9,14,18-tetrone

EC Number:

201-375-5

EC Name:

6,15-dihydroanthrazine-5,9,14,18-tetrone

Cas Number:

81-77-6

Molecular formula:

C28H14N2O4

IUPAC Name:

6,15-dihydroanthrazine-5,9,14,18-tetrone

Details on test material:

- Purity: Lab sample
- Storage: room temperature
- Physical state: solid, insoluble in water
- Stability: guaranteed by sponsor

Method

Target gene:

Histidine operon

Species / strain

Species / strain / cell type:

S. typhimurium TA 1535, TA 1537, TA 98 and TA 100

Metabolic activation:

with and without

Metabolic activation system:

Aroclor 1254 induced rat liver S-9 fraction mixed with a series of cofactors

Test concentrations with justification for top dose:

20, 78, 313, 1,250 and 5000 µg/0.1 mL

Vehicle / solvent:

- Vehicle: DMSO

Controls

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: without metabolic activation: Daunorubincin-HCl, 4-Nitroquinoline-N-Oxide, Sodium azide, 9(5) aminoacridine HCl; with metabolic activation: 2 aminoanthracene and cyclophosphamide

Remarks:

further information of positive control substances; see "any other information on materials and methods"

Details on test system and experimental conditions:

METHOD OF APPLICATION: in agar (plate incorporation test)

DURATION
- Exposure duration: 48 hours at 37°C

NUMBER OF REPLICATIONS: 2 experiments; 3 cultures / experiment

DETERMINATION OF CYTOTOXICITY
- Method: Reduction of revertants or reduction or clearing of background lawn

Evaluation criteria:

Criteria for a positive response:
The test substance is considered to be positive in this test system if one or both of the following conditions are met:
- a reproducible doubling of the mean number of revertants per plate above that of the negative control at any concentration level for one or more of the following strains: TA 98, TA 1535 and TA 1537,
- a reproducible increase of the mean number of revertants per plate for any concentration above that of the negative control by a factor of 1.5 for strain TA 100. Generally a concentration-related effect should be demonstrable.

Assay acceptance criteria:
A test is considered acceptable if the mean colony counts of the control values of all strains are within the historical control ranges and if the results of the positive controls meet the criteria for a positive response.

Acceptable range for negative control (without metabolic activation).
Revertants/plate
TA 98: 12-50
TA 100: 80-220
TA 1535: 7-30
TA 1537: 3-20

Acceptable range for negative control (with metabolic activation).
Revertants/plate
TA 98: 20-70
TA 100: 70-220
TA 1535: 7-35
TA 1537: 5-30

Statistics:

Arithmetical mean was calculated of the number of revertants.

Results and discussion

Test results

Species / strain:

S. typhimurium TA 1535, TA 1537, TA 98 and TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity, but tested up to precipitating concentrations

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
- Precipitation: Precipitation of the test item in soft agar was observed at concentrations of 313 µg/0.1 mL and above.

RANGE-FINDING/SCREENING STUDIES:
- Strain tested: TA 100
- Metabolic activation: without
- Concentrations: 0.08, 0.31, 1.2, 4.9, 19.5, 78.1, 312.5, 1250.0, 5000 µg/0.1 mL
- Repetitions: 2 cultures/concentration
- Method: the protocol used was the same as in the mutagenicity test
- Results: 5000 µg/0.1 mL of the test substance did not lead to a growth-inhibiting effect. This concentration was therefore selected as the highest in the mutagenicity tests. Number of revertants at 5000 µg/0.1 mL = 132 vs. 99 in control.

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'.

Any other information on results incl. tables

Table 2: Experiment 1: Mean values (without S9 mix)

			TA 98		TA 100		TA 1535		TA 1537
	Concentration (µg/0.1 ml)	S9	X (n=3)	revertant factor	X (n=3)	revertant factor	X (n=3)	revertant factor	X (n=3)	revertant factor
Test substance	control	no	27		144		12		10
	20	no	18	0.7	127	0.9	11	0.9	6	0.6
	78	no	21	0.8	140	1.0	11	0.9	7	0.7
	313*	no	21	0.8	139	1.0	13	1.1	8	0.8
	1250*	no	22	0.8	145	1.0	12	1.0	11	1.1
	5000*	no	30	1.1	144	1.0	11	0.9	11	1.1
positive control	control	no	19		131		10		6
	Low conc.	no	681	35.8	837	6.4	776	7.8	105	17.8
	High conc.	no	881	46.4	1447	11.0	1155	115.5	1155	192.5

 

Table 3: Experiment 1: Mean values (with S9 mix)

			TA 98		TA 100		TA 1535		TA 1537
	Concentration (µg/0.1 ml)	S9	X (n=3)	revertant factor	X (n=3)	revertant factor	X (n=3)	revertant factor	X (n=3)	revertant factor
Test substance	control	yes	31		125		16		15
	20	yes	36	1.2	128	1.0	24	1.5	9	0.6
	78	yes	31	1.0	139	1.1	16	1.0	13	0.9
	313*	yes	36	1.2	149	1.2	17	1.1	8	0.5
	1250*	yes	51	1.6	129	1.0	9	0.6	13	0.9
	5000*	yes	37	1.2	143	1.1	8	0.5	20	1.3
positive control	control	yes	32		163		14		18
	Test conc.	yes	1246	38.9	1611	9.9	530	37.9	291	16.2

 

Table 4: Experiment 2: Mean values (without S9 mix)

			TA 98		TA 100		TA 1535		TA 1537
	Concentration (µg/plate)	S9	X (n=4)	revertant factor	X (n=4)	revertant factor	X (n=4)	revertant factor	X (n=4)	revertant factor
Test substance	control	no	17		146		12		7
	20	no	18	1.0	179	1.2	10	0.9	7	1.0
	78	no	17	1.0	191	1.3	7	0.6	6	0.9
	313*	no	19	1.1	165	1.1	8	0.7	9	1.4
	1250*	no	21	1.2	147	1.0	13	1.1	7	1.0
	5000*	no	21	1.2	161	1.1	10	0.8	9	1.3
Positive control	Control	no	11		138		9		5
	Low conc.	no	295	17.7	263	1.9	523	58.1	49	9.8
	High conc.	no	752	68.4	496	3.4	911	101.2	1048	209.6

 

Table 5: Experiment 2: Mean values (with S9 mix)

			TA 98		TA 100		TA 1535		TA 1537
	Concentration (µg/plate)	S9	X (n=4)	revertant factor	X (n=4)	revertant factor	X (n=4)	revertant factor	X (n=4)	revertant factor
Test substance	control	yes	29		136		13		10
	20	yes	23	0.8	137	1.0	15	1.1	10	1.0
	78	yes	29	1.0	164	1.2	9	0.7	9	0.9
	313*	yes	30	1.0	164	1.2	10	0.8	11	1.1
	1250*	yes	29	1.0	169	1.2	9	0.7	11	1.1
	5000*	yes	36	1.3	180	1.3	10	0.8	9	0.9
Positive control	Control	no	29		127		11		6
	Test conc.	no	1288	44.4	965	7.6	502	45.6	109	18.2

 

  * Precipitation on test plate

 

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information):
negative