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Toxicological information

Genetic toxicity: in vitro

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Administrative data

Endpoint:
in vitro gene mutation study in bacteria
Type of information:
experimental study
Adequacy of study:
weight of evidence
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
data from handbook or collection of data
Justification for type of information:
Data is from peer reviewed publication

Data source

Reference
Reference Type:
publication
Title:
Mutagenicity of mono-nitrobenzene derivatives in the Ames test and rec assay
Author:
Makoto Shimizu and Eiji Yan
Year:
1986
Bibliographic source:
Mutation Research, 170 (1986) 11-22

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
other: Refer below principle
Principles of method if other than guideline:
Gene mutation toxicity study was performed to evaluate the mutagenic nature of p-Nitrobenzonitrile
GLP compliance:
not specified
Type of assay:
bacterial reverse mutation assay

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material: p-Nitrobenzonitrile
- IUPAC name: 4-Nitrobenzonitrile
- Molecular formula: C7H4N2O2
- Molecular weight: 148.121 g/mol
- Substance type: Organic
Specific details on test material used for the study:
- Name of test material: p-Nitrobenzonitrile
- IUPAC name: 4-Nitrobenzonitrile
- Molecular formula: C7H4N2O2
- Molecular weight: 148.121 g/mol
- Substance type: Organic
- Physical state: No data
- Purity: 98% min
- Impurities (identity and concentrations): 2%

Method

Target gene:
Histidine
Species / strain
Species / strain:
other: TA98, TA100, TA1535, TA1538 and TA1537
Details on mammalian cell lines (if applicable):
Not applicable
Additional strain characteristics:
not specified
Cytokinesis block (if used):
No data
Metabolic activation:
with and without
Metabolic activation system:
The S-9 fraction was obtained from PCB-induced hamster or rat liver
Test concentrations with justification for top dose:
0, 0.01, 0.05, 0.1, 0.5, 1.0 or 5.0 mg/plate
Vehicle:
- Vehicle(s)/solvent(s) used: DMSO
- Justification for choice of solvent/vehicle: The chemical was soluble in DMSO
Controls
Negative controls:
not specified
Solvent controls:
yes
Remarks:
DMSO
True negative controls:
not specified
Positive controls:
yes
Positive control substance:
9-aminoacridine
2-nitrofluorene
N-ethyl-N-nitro-N-nitrosoguanidine
other: 2-aminoanthracene (2-AA) (S9 mix added only)
Details on test system and conditions:
METHOD OF APPLICATION: preincubation

DURATION
- Preincubation period: 15 mins
- Exposure duration: 70 hrs in dark
- Expression time (cells in growth medium): 70 hrs in dark
- Selection time (if incubation with a selection agent): No data
- Fixation time (start of exposure up to fixation or harvest of cells): No data

SELECTION AGENT (mutation assays): No data
SPINDLE INHIBITOR (cytogenetic assays): No data
STAIN (for cytogenetic assays): No data

NUMBER OF REPLICATIONS: All tests were performed in duplicate and repeated at least 3 times separately

NUMBER OF CELLS EVALUATED: No data

DETERMINATION OF CYTOTOXICITY
- Method: mitotic index; cloning efficiency; relative total growth; other: Yes, The background bacterial lawn was routinely checked by microscopy, as high doses of the complexes proved toxic to all strains resulting in a thinning out of the bacterial lawn.

OTHER EXAMINATIONS:
- Determination of polyploidy: No data
- Determination of endoreplication: No data
- Other: No data

OTHER: First the tests were carried out without metabolic activation and were terminated if the mutagenicity was positive. In case of negative results, tests with metabolic activation were carried out in addition.

Two strains (TA98 and TA100) were checked routinely for the presence of the ampicillin resistance for the R factor.
Rationale for test conditions:
No data
Evaluation criteria:
Chemicals inducing more than twice the number of revertant colonies on the control plate were considered as mutagenic.
Statistics:
No data

Results and discussion

Test results
Species / strain:
other: TA98, TA100, TA1535, TA1538 and TA1537
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity:
yes
Remarks:
At 10 mg/plate
Vehicle controls valid:
yes
Negative controls valid:
not specified
Positive controls valid:
yes
Remarks on result:
other: No mutagenic potential
Additional information on results:
No data

Any other information on results incl. tables

Table: Mutagenicity of p-Nitrobenzonitrile

Dose

Revertants/plate

TA98

TA1538

TA1537

TA100

TA1535

-

+

-

+

-

+

-

+

-

+

0.01

28±4

28±3

24±3

26±3

9±2

10±2

206±19

189±17

35±6

30±5

0.05

26±3

30±4

28±3

24±3

10±2

8±1

180±18

201±23

30±5

34±3

0.1

40±5

30±4

26±3

25±3

11±3

10±1

199±21

224±32

30±4

32±4

0.5

35±3

33±4

30±3

26±4

8±2

11±2

198±23

216±28

21±3

29±5

1

8±7

20±3

0*

12±6*

0*

0*

0*

190±3

0*

9±8*

5

0*

0*

0*

0*

0*

0*

0*

0*

0*

0*

*: Indicates toxic effects

 

Applicant's summary and conclusion

Conclusions:
p-Nitrobenzonitrile did not induce mutation in Salmonella typhimurium strains TA98, TA100, TA1535, TA1538 and TA1537 in the presence and absence of S9 metabolic activation system and hence is not likely to classify as a gene mutant in vitro.
Executive summary:

Gene mutation toxicity study was performed to determine the mutagenic nature of p-Nitrobenzonitrile. The study was performed using Salmonella typhimurium strains TA98, TA100, TA1535, TA1538 and TA1537 in the presence and absence of S9 metabolic activation system using the preincubation protocol. The chemical was dissolved in DMSO as solvent and used at dose levels 0, 0.01, 0.05, 0.1, 0.5, 1.0 or 5.0 mg/plate by the preincubation for 15 mins. The plates were incubated for 70 hrs in dark and observed for the presence of colonies. Concurrent solvent and positive control chemicals were also included in the study. p-Nitrobenzonitriledid not induce mutation inSalmonella typhimurium strains TA98, TA100, TA1535, TA1538 and TA1537 in the presence and absence of S9 metabolic activation system and hence is not likely to classify as a gene mutant in vitro.