p-anisidine

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:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Data is from authoritative source

Data source

Reference

Reference Type:

publication

Title:

Enhanced mutagenicity of anisidine isomers in bacterial strains containing elevated N-acetyltransferase activity

Author:

David C. Thompson, P. David Josephy, Joseph W.K. Chu and Thomas E. Eling

Year:

1992

Bibliographic source:

Mutation Research. 279 (1992) 83-89

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

GLP compliance:

not specified

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

p- anisidine

IUPAC Name:

p- anisidine

Constituent 2

Reference substance name:

p-anisidine

EC Number:

203-254-2

EC Name:

p-anisidine

Cas Number:

104-94-9

Molecular formula:

C7H9NO

IUPAC Name:

4-methoxyaniline

Details on test material:

Name of test material (as cited in study report): p- anisidine
- Molecular formula (if other than submission substance): C7-H9-N-O
- Molecular weight (if other than submission substance): 123.1541
- Substance type: Organic
- Physical state: Solid
- Purity: No data available
- Impurities (identity and concentrations): No data available

Method

Target gene:

hisG46

Species / strain

Species / strain / cell type:

S. typhimurium, other: YG1029

Additional strain / cell type characteristics:

acetyltransferase proficient

Remarks:

YG1029 (TA1OO pYG 219)

Metabolic activation:

with and without

Metabolic activation system:

Ram seminal vesicle microsomes (RSVM) Hamster S9 and horseradish peroxidase

Test concentrations with justification for top dose:

Highest dose 10 µmoles/plate

Vehicle / solvent:

DMSO

Controls

Untreated negative controls:

not specified

Negative solvent / vehicle controls:

not specified

True negative controls:

not specified

Positive controls:

not specified

Positive control substance:

not specified

Details on test system and experimental conditions:

METHOD OF APPLICATION: in agar (plate incorporation)

DURATION
- Preincubation period: For RSVM activated mutagenicity): Mutagen, RSVM in buffer and bacterial culture were mixed and incubated initially for 3 mins and after addition of arachidonic acid incubation was further continued for 30 mins

For S9 activated mutagenicity: Mutagen, S9 in buffer and bacterial culture were mixed and incubated for 30 mins

For Horse radish peroxidase/hydrogen peroxide system: No data available

- Exposure duration: No data available
- Expression time (cells in growth medium): about 14 hrs
- Selection time (if incubation with a selection agent): No data available
- Fixation time (start of exposure up to fixation or harvest of cells): No data available

SELECTION AGENT (mutation assays): Tetracycline (6.25 µg/ml)

SPINDLE INHIBITOR (cytogenetic assays): No data available

STAIN (for cytogenetic assays): No data available

NUMBER OF REPLICATIONS: No data available

NUMBER OF CELLS EVALUATED: No data available

DETERMINATION OF CYTOTOXICITY
- Method: No data available

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

OTHER: Overnight cultures were grown at 37°C with shaking at 120 rpm in Oxoid Nutrient Broth No. 2 for about 14 hrs, until the OD, reached 1.0.

Statistics:

Colonies were counted after 48 h, using a 3M 620 automatic colony counter with a size threshold of 0.29 mm.

Doses are plotted using the transformed variable X = (1 + dose) in logarithmic scale; this method allows the zero dose point to be included without breaking the axis.

Results and discussion

Test results

Species / strain:

S. typhimurium, other: YG1029

Metabolic activation:

with and without

Genotoxicity:

positive

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

not specified

Untreated negative controls validity:

not specified

Positive controls validity:

not specified

Additional information on results:

For YG1029:
With metabolic activation:
RSVM mutagenicity system: Positive
S9 mutagenicity system: Positive
Horse radish peroxidase/hydrogen peroxide system: Positive (at the two highest concentrations hydrogen peroxide examined i.e. 0.1 and 0.2 µmoles/ plate)

Without metabolic activation:
RSVM mutagenicity system: Positive (very little mutagenicity)
S9 mutagenicity system: Positive (very little mutagenicity)
Horse radish peroxidase/hydrogen peroxide system: Positive (at the two highest concentrations of hydrogen peroxide examined i.e. 0.1 and 0.2 µmoles/ plate)

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'.

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information):
positive

Mutagenic response of p- anisidine was observed for Salmonella typhimurium tester strain YG1029 with and without metabolic activation using RSVM, hamster S9 and horse radish peroxidase/hydrogen peroxide system. Genetic toxicity results were observed to be positive in this study.

Executive summary:

New bacterial tester strains ofSalmonella typhimuriumhave been constructed which have greatly elevated sensitivities to aromatic amines and nitroaromatic compounds; these strains carry plasmid-borne copies of the genes encoding the bacterial enzymes acetyl CoA-dependent arylamine N-acetyltransferase/ arylhydroxylamine O-acetyltransferase (NAT/OAT) or nitroreductase.

 

One of these new strains includes YG1029 (derived from TA100). Mutagenic response of p-anisidine in the Ames test using strain YG1029 has been tested in the given study.

 

Metabolic activation have been carried out using Hamster S9 liver homogenate or ram seminal vesicle microsomes as activating systems and the possible role of N-acetyltransferase enzymes in the mutagenicity and carcinogenicity of p-anisidine has been studied.

 

Horseradish peroxidase as a possible activating system for p-anisidine in YG1029 has also been studied. It catalyzes the oxidation of xenobiotics such as aromatic amines to free radical and other reactive intermediates.

When tested with YG1029, however, p-anisidine caused a dose-dependent increase in revertants. p-Anisidine was mutagenic with either hamster S9 or RSVM as an activating system. The number of p-anisidine-dependent revertants observed with the two activating systems (S9, RSVM) was similar.

 

With RSVM, the presence or absence of arachidonic acid had no effect on the number of revertants.

 

The horseradish peroxidase/ hydrogen peroxide system was unable to activate p-anisidine to a mutagen. At the two highest doses of hydrogen peroxide examined (0.1 and 0.2 µmoles/ plate), some toxicity was evident.

 

The above study suggests p-anisidine to be mutagenic for the Salmonella typhimurium tester strain YG1029.