Pyrimido[5,4-g]pteridine-2,4,6,8-tetramine, 4-methylbenzenesulfonate, base-hydrolyzed

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:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP, guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2001

Report date:

2001

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

-

EC Number:

440-560-7

EC Name:

-

Cas Number:

346709-25-9

Molecular formula:

Unspecified

IUPAC Name:

2,6,8-triamino-4H,10H-pyrimido[5,4-g]pteridin-4-one; pyrimido[5,4-g]pteridine-2,4,6,8-tetramine

Details on test material:

- Description: Yellow powder
- Purity: 94.8 %
- Test substance storage: At room temperature in the dark
- Stability under storage conditions: Stable
- Expiry date: 06 November 2005
- Stability in vehicle - Dimethyl sulfoxide: at least 4 h

Method

Target gene:

His: Salmonella
Trp: E. Coli

Species / strain

Species / strain / cell type:

other: Salmonella typhimurium strains: TA100, TA98, TA1535, TA1537; E. coli : WP2uvrA

Metabolic activation:

with and without

Metabolic activation system:

Aroclor 1254 induced rat liver S9 mix

Test concentrations with justification for top dose:

first mutation assay (range finding test): 10, 33, 100, 333, 1000, 3330 and 5000 µg/plate
second mutation assay: 10, 33, 100, 333 and 1000 µg/plate

Vehicle / solvent:

Dimethyl sulfoxide

Controls

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: see: "Details on test system and conditions"

Details on test system and experimental conditions:

POSITIVE CONTROLS:
Without metabolic activation (-S9-mix):
TA1535: sodium azide (SA), 5 µg
TA1537: 9-aminoacridine (9AC), 60 µg
TA98: daunomycine (DM), 4 µg
TA100: methylmethanesulfonate (MMS), 650 µg
WP2uvrA: 4-nitroquinoline N-oxide (4-NQO), 10 µg

With metabolic activation (+S9-mix):
TA1537: 2-aminoanthracene (2AA), 2.5 µg
TA1535,TA98 and TA100: 2-aminoanthracene (2AA), 1 µg
WP2uvrA: 2-aminoanthracene (2AA), 5 µg; In the presence of 10% (v/v) S9-fraction, the concentration of 2AA was 10 µg/plate

METHOD OF APPLICATION: in agar (plate incorporation)

DURATION
- Incubation period: 48 hours. After this period revertant colonies (histidine independent for Salmonella typhimurium bacteria and tryptophan independent for Escherichia coli) were counted.
- COLONY COUNTING:
- Exposure duration: The revertant colonies (histidine independent/ tryptophan independent) were counted automatically with a Protos model 50000 colony counter or manually, if less than 40 colonies per plate were present. Plates with sufficient test article precipitate to interfere with automated colony counting were counted manually.

NUMBER OF REPLICATIONS: 3

Evaluation criteria:

A test substance is considered negative (not mutagenic) in the test if:
a) 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.
b) The negative response should be reproducible in at least one independently repeated experiment.

A test substance is considered positive (mutagenic) in the test if:
a) 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.
b) The positive response should be reproducible in at least one independently repeated experiment.
The preceding criteria were not absolute and other modifying factors might enter into the final evaluation decision.

Results and discussion

Test results

Species / strain:

other: Salmonella typhimurium strains: TA100, TA98, TA1535, TA1537; E. coli : WP2uvrA

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 applicable

Positive controls validity:

valid

Additional information on results:

PRECIPITATE: The test substance precipitated in the top agar at concentrations of 100 µg/plate and upwards. Precipitation of the test substance on the plates was observed at the start and at the end of the incubation period at concentrations of 1000 µg/plate and upwards in all tester strains.

TOXICITY: No reduction of the bacterial background lawn and no biologically significant decrease in the number of revertants were observed.

MUTAGENICITY: No increase in the number of revertants was observed upon treatment with the test substance under all conditions tested (table 1). All bacterial strains showed negative responses over the entire dose range, i.e. no dose-related, two-fold, increase in the number of revertants in two independently repeated experiments.

Any other information on results incl. tables

Table 1: Number of revertants in the control or after treatment with the test substance

First experiment (10 - 5000 µg/plate)
Strain	Metabolic activation system	mean revertants in Controls	maximum revertant factor	dose dependency	Assessment
TA 1535	no	11	1.27	no	negative
	yes	14	1.28	no	negative
TA 1537	no	12	0.67	no	negative
	yes	8	1.13	no	negative
TA 98	no	15	1.20	no	negative
	yes	22	1.27	no	negative
TA 100	no	85	1.24	no	negative
	yes	98	1.05	no	negative
E. coli WP2 uvrA	no	12	1.08	no	negative
	yes	10	1.40	no	negative
Second experiment (10 - 1000 µg/plate)
Strain	Metabolic activation system	mean revertants in Controls	maximum revertant factor	dose dependency	Assessment
TA 1535	no	18	1.11	no	negative
	yes	14	1.57	no	negative
TA 1537	no	3	1.67	no	negative
	yes	3	1.67	no	negative
TA 98	no	19	1.05	no	negative
	yes	19	1.11	no	negative
TA 100	no	68	1.16	no	negative
	yes	65	1.15	no	negative
E. coli WP2 uvrA	no	12	1.16	no	negative
	yes	13	1.38	no	negative