[1,1'-Bianthracene]-9,9',10,10'-tetrone, 4,4'-bis[[4,6-bis[[3-(diethylamino)propyl]amino]-1,3,5-triazin-2-yl]amino]-

Genetic toxicity: in vivo

Administrative data

Endpoint:

in vivo mammalian somatic cell study: cytogenicity / erythrocyte micronucleus

Remarks:

Type of genotoxicity: chromosome aberration

Type of information:

experimental study

Adequacy of study:

key study

Study period:

14 April 1993 to 6 July 1993

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Study conducted according to EU Method B.12 under GLP

Cross-reference

Cross-reference 1

Reason / purpose for cross-reference:

reference to same study

Cross-reference 2

Reason / purpose for cross-reference:

reference to other study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1993

Report date:

1993

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

EU Method B.12 (Mutagenicity - In Vivo Mammalian Erythrocyte Micronucleus Test)

Deviations:

yes

Remarks:

only male animals were used

Principles of method if other than guideline:

Not relevant

GLP compliance:

yes (incl. QA statement)

Type of assay:

micronucleus assay

Test material

Test material information

Constituent 1

Reference substance name:

308 AQN

IUPAC Name:

308 AQN

Test material form:

solid: particulate/powder

Remarks:

migrated information: powder

Test animals

Species:

mouse

Strain:

CD-1

Sex:

male

Details on test animals or test system and environmental conditions:

Sufficient male albino CD1 strain mice were supplied by Charles River (UK) Ltd., Manston, Kent. At the start of the main study the mice weighed 24 - 31g and were approximately five to seven weeks old. After a minimum acclimatisation period of five days the animals were selected at random and given a unique number within the study by ear punching and a number written on a colour cage card.

Administration / exposure

Route of administration:

intraperitoneal

Vehicle:

Suspension in arachis oil B.P.

Details on exposure:

Dose Level:
2000 mg/kg oral
2000, 1000, 500, 250, 125, 60, 30, 20, 10, 5 mg/kg i.P.

Duration of treatment / exposure:

all animals were dosed once

Frequency of treatment:

once

Post exposure period:

observation period 2 days

Doses / concentrations

Doses / concentrations 1

Remarks:

Doses / Concentrations:
20 mg/kg
Basis:
nominal conc.

Doses / concentrations 2

Remarks:

Doses / Concentrations:
10 mg/kg
Basis:
nominal conc.

Doses / concentrations 3

Remarks:

Doses / Concentrations:
5 mg/kg
Basis:
nominal conc.

No. of animals per sex per dose:

Groups of seven male mice per dose were used.
In addition three further groups of male mice were included in the study; two groups of seven mice were dosed wia the intraperitoneal route with the vehicle alone (arachis oil B.P.) and the third group (five mice) was dosed orally with cyclophosphamide, a positive control material.

Control animals:

yes

Positive control(s):

One group (five mice) was dosed orally with cyclophosphamide, a positive control material.

Examinations

Tissues and cell types examined:

Both femurs of each test animals were examined.

Details of tissue and slide preparation:

Immediately following sacrifice, both femurs were dissected from each animal, aspirated with foetal calf serum and bone marrow smears prepared following centrifugation and re-suspension. The smears were air-dried, fixed in absolute methanol, and stained in May-Grünwald/Giemsa.

Evaluation criteria:

Stained bone marrow smears were coded and eamined blind using light microscopy at x 1000 magnification. the incidence of micronucleated cells per 2000 polychromatic erythrocytes (PCE-blue stained immature cells) per animal was scored, in this study the incidence of micronucleated cells per 4000 polychromatic erythrocytes was scored for the 24 hour 20 mg/kg group due to only four animals being available for evaluation and consequently statistical analysis. Micronuclei are normally circular in shape, although occassionally they may be oval or half-moon shaped, and have a sharp contour with even staining. In addition, the number of normochromatic erythrocytes (NCE-pink stained mature cells) associated with 1000 total erythrocytes were counted; these cells were also scored for incidence of micronuclei.

When a micronucleus slide is scored the slide assessor scores NCE and PCE until a total of 1000 erythrocytes (NCE + PCE) have been scored. At this point the number of NCE's is recorded and no more NCE's are scored. The slide scorer then continues scoring PCE's until a total of 2000 PCE's have been scored. Therefore the PCE/NCE ratio is obtained during the first half of 2000 scored PCE.

PCE number scored is usually 2000 unless, due to sample loss or insufficient bone marrow obtained, fewer PCE's are scored. PCE + MN relates to the number of PCE's with micronuclei seen within the number of PCE's scored. NCE number scored relates to the number of NCE scored within 1000 total erythrocytes (PCE + NCE), and NCE + MN relates to the number of NCE seen within this number with micronuclei. PCE/NCE ratio is obtained from the equation (1000 -NCE scored)/NCE scored.

Statistics:

The individual ratio of polychromatic to normochromatic erythrocytes was calculated together with appropriate group mean values and standard deviations.

Results and discussion

Test results

Sex:

male

Genotoxicity:

negative

Vehicle controls validity:

valid

Negative controls validity:

not applicable

Positive controls validity:

valid

Additional information on results:

There were premature deaths in animals dosed via theintraperitoneal route with 308 AQN at and above 20 mg/kg.

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information): negative
308 AQN was considered to be non-genotoxic under the conditions of the test.

Executive summary:

There was no evidence of toxicologically significant increases in the incidence of micronucleated polychromatic erythrocytes in animals dosed with 308 AQN when compared to the concurrent vehicle control groups. No significant change in the PCE/NCE ratio was observed after dosing with 308 AQN.

The positive control material produced a large and significant increase in the frequency of micronucleated polychromatic erythrocytes.

The test material, 308 AQN, was considered to be non-genotoxic under the conditions of the test.