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Toxicological information

Genetic toxicity: in vivo

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Administrative data

Endpoint:
in vivo mammalian cell study: DNA damage and/or repair
Remarks:
Type of genotoxicity: DNA damage and/or repair
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Comparable to guideline study with acceptable restrictions (limited documentation)

Data source

Reference
Reference Type:
publication
Title:
Unnamed
Year:
1982

Materials and methods

Test guideline
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 486 (Unscheduled DNA Synthesis (UDS) Test with Mammalian Liver Cells in vivo)
Deviations:
yes
Remarks:
only one sampling time (12 h) and only 50 cells/animal evaluated
GLP compliance:
not specified
Type of assay:
unscheduled DNA synthesis

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): nitrobenzene; Eastman Kodak
- Analytical purity: no data

Test animals

Species:
rat
Strain:
Fischer 344
Sex:
male
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Breeding Labs, Kingston, NY
- Weight at study initiation: 175 - 275 g
- Diet (e.g. ad libitum): NIH-07 feed (Ziegler Brothers, Gardner, PA), ad libitum
- Water (e.g. ad libitum): untreated tap water, ad libitum

Sera from the animals were tested and did not contain antibodies against pneumonia virus of mice, revirus 3, GD VII, Kilham rat virus, H-1, Sendai, mouse adenovirus, mouse hepatitis virus, lymphocytic chorioimeningitis virus or rat corona virus.

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
- Vehicle(s)/solvent(s) used: corn oil
- Amount of vehicle (if gavage or dermal): 0.1 - 0.4 mL/100g body weight
Details on exposure:
The highest dose was generally selected to be at or near the LD50 when such information is available.
Duration of treatment / exposure:
single dose
Post exposure period:
Hepatocytes were isolated 12 h after treatment.
Doses / concentrations
Remarks:
Doses / Concentrations:
200 and 500 mg/kg in corn oil
Basis:
actual ingested
No. of animals per sex per dose:
3
Control animals:
yes, concurrent vehicle
Positive control(s):
2-acetylaminofluorene; methylmethanesulfonate, benzidine (genotoxic hepatocarcinogens)
- Route of administration: oral, gavage
- Doses / concentrations: 2-AAF: 50 mg/kg; MMS: 100 mg/kg; benzidine: 200 mg/kg

Examinations

Tissues and cell types examined:
freshly isolated hepatocytes
Details of tissue and slide preparation:
CRITERIA FOR DOSE SELECTION: The highest dose was generally selected to be at or near the LD50 when such information is available.


DETAILS OF SLIDE PREPARATION: Cultures were washed twice with Williams Medium E (WE), fixed followed by six washes with deionized water. When dry, coverslips were mounted with permount and dipped in Kodak NTB-2 emulsion diluted 1.1 with water. Slides were exposed for 12 - 14 days at -20°C and developed for quantitative autoradiography.


METHOD OF ANALYSIS: An area of the slide was randomly selected and 50 morphologically unaltered cells were counted (ARKTEK Model 880 colony counter interfaced to a microscope). All counts were made on the "OBJ AREA" mode. The highest of three nuclear-sized areas over the cytoplasm and adjacent to the nucleus was subtracted from the nuclear count to give net grains/nucleus. The percentage of cells in repair, which is a useful indicator of the extent of damage throughout the liver, is defined as those exhibiting five or more net grains.


OTHER: Cells were incubated in WE medium containing 10% fetal calf serum for ca. 90 min at 37 °C to allow attachment of the cells to the coverslips. Cultures were washed and incubated in WE containing 10 µCi/mL 3H-thymidine for 4 hours. Cultures were again washed and incubated overnight (14 - 16 h) in 0.25 mM thymidine.
Evaluation criteria:
Five or more net grains/nucleus was considered positive.

Results and discussion

Test results
Sex:
male
Genotoxicity:
negative
Toxicity:
not specified
Vehicle controls validity:
valid
Negative controls validity:
not examined
Positive controls validity:
valid

Any other information on results incl. tables

Chemical

Dose [mg/kg]

Time [h]

Net grains ± SE

Cells in repair [%] ± SE

Corn oil

2

-5.1 ± 0.5

1 ± 0

12

-4.4 ± 0.5

3 ± 1

Nitrobenzene

200

12

-4.4 ± 1.2

5 ± 2

500

12

-5.6 ± 1.3

6 ± 3

2-AAF

50

2

13.2 ± 2.7

71 ± 7

12

45.0 ± 11.3

96 ± 3

Applicant's summary and conclusion