3,7-dinitroso-1,3,5,7-tetraazabicyclo[3.3.1]nonane

Genetic toxicity: in vivo

Administrative data

Endpoint:

in vivo mammalian somatic cell study: cytogenicity / erythrocyte micronucleus

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Data source

Reference

Reference Type:

publication

Title:

Evaluation of some formaldehyde-release compounds and other biocides in the mouse micronucleus test

Author:

C.R. Richardson, J.A. Styles and B. Burlinson

Year:

1983

Bibliographic source:

Mutation Research, 124 (1983) 241-246 (Elsevier)

Materials and methods

GLP compliance:

not specified

Type of assay:

mammalian erythrocyte micronucleus test

Test material

Test material information

Constituent 1

Reference substance name:

3,7-dinitroso-1,3,5,7-tetraazabicyclo[3.3.1]nonane

EC Number:

202-928-3

EC Name:

3,7-dinitroso-1,3,5,7-tetraazabicyclo[3.3.1]nonane

Cas Number:

101-25-7

Molecular formula:

C5H10N6O2

IUPAC Name:

3,7-dinitroso-1,3,5,7-tetraazabicyclo[3.3.1]nonane

Test material form:

solid

Details on test material:

- Chemical name: Dinitrosopentamethylenetetramine
- CAS: 101-25-7
- EC: 202-928-3
- MW: 186.17 g/mol
- pale yellow crystalline powder

Specific details on test material used for the study:

DNPT supplied by Organics Division, ICI PLC, Blackley, Manchester, Great Britain

Test animals

Species:

mouse

Strain:

C57BL

Remarks:

/6J

Sex:

male

Details on test animals or test system and environmental conditions:

ANIMALS
Male C57Bl/6J mice (6-8 weeks old) were used for both the main study and the LD50 determinations.

HUSBANDRY
On arrival the mice were housed 5 per cage on mobile mouse racks and given food (Porton Combined Diet [PCD] supplied by BP Nutrition Ltd., Stepfield, Witham, Essex, Great Britain) and water ad libitum.
The animal cell was maintained at a mean temperature of 21°C (range 20-22°C), a relative humidity mean of 44% (range 29-58%) with 22 air changes/h.
Lighting was cycled with 12 h light and 12 h dark each 24 h.

Administration / exposure

Route of administration:

intraperitoneal

Vehicle:

not specified

Details on exposure:

Animals were dosed with 2 consecutive daily doses administered 24 h apart using doses equivalent to 80% and 50% of the LD50/7 by intraperitoneal injection.
The lower dose level was used to ensure a test result in the event of deaths in the top-dose group and to provide additional data in the event of a positive result.

Duration of treatment / exposure:

24 h

Frequency of treatment:

2 consecutive daily doses administered 24 h apart

Post exposure period:

24-48 h

Doses / concentrations

Dose / conc.:

160 other: mg/kg

No. of animals per sex per dose:

- Negative control a: 10 (24h) / 11 (48h)
- DNPT: 7 (24h) / 6 (48h)
- Positive control: 9 (48h)

Control animals:

yes

Positive control(s):

Cyclophosphamide (75 mg/kg)

Examinations

Tissues and cell types examined:

polychromtic erythrocytes (PCEs) from bone marrow (femur)

Details of tissue and slide preparation:

The animals were killed by cervical dislocation either 24 or 48 h after the last dose of compound. Femurs were removed and stripped clean of muscle. The iliac end of the femur was removed and a fine paint brush wetted with a solution of albumen (6% w/v in water) was dipped into the marrow canal. 3-4 streaks of marrow suspension were then applied to appropriately labelled clean dry microscope slides.
The slides were allowed to air-dry and then stained in Leishman's stain using an Ames Hema-Tek staining machine (Miles Laboratories Inc., Ames Division, Stoke Court, Stoke Poges, Slough, Bucks., Great Britain). Slides were coded and scored blind: 500 polychromatic erythrocytes (PCEs) were examined for each animal and the number containing micronuclei scored. The ratio of polychromatic to mature erythrocytes was also determined as a measure of cytotoxicity.
Slides prepared from animals dosed at 50% of the LD50/7 were removed before coding. In the event of a positive result being observed with any of the test chemicals, the relevant slides from the low-dose groups were randomised with positive and negative control slides and scored for the presence of micronuclei.

Statistics:

The results were analysed for significant differences from the control groups using a one-sided Student's ' t ' test.

Results and discussion

Test results

Key result

Sex:

male

Genotoxicity:

negative

Toxicity:

yes

Remarks:

80% of the LD50 based on preliminary toxicity test

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

DNPT gave negative results at both the sampling times used.
The system positive control (cyclophosphamide) gave a greatly elevated and statistically significant increase in micronuclei.

Any other information on results incl. tables

Incidence of micronuclei/1000 cells (2 sampling times)

Substance	Dose	24h sampling	48h sampling
NC (Control a)		1.6 +/- 2.1	1.4 +/- 1.6
DNPT	160 mg/kg	1.4 +/- 1.5	1.3 +/- 1
PC (Cyclophosphamide)	75 mg/kg	-	64.1 +/- 18

There was no undue influence of cytotoxicity on the results generated, since the ratios of PCEs to mature erythrocytes did not deviate from the normally expected range.

Applicant's summary and conclusion

Conclusions:

Under the conditions of this test, the test item did not show clastogenic activity.

Executive summary:

A number of biocidal chemicals were tested for clastogenic activity in the micronucleus test using C57Bl/6J mice.

N, N-dinitrosopentamethylenetetramine (DNPT) was tested as the negative control.

DNPT did not increase the incidence of micronuclei.