Adipic acid

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

1996

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

test procedure in accordance with national standard methods

Data source

Reference

Reference Type:

publication

Title:

Mutagenicity test data of existing chemical substances

Author:

Chemical Substance Investigation Division, Industrial Safety and Health Department, Labour Standatds Bureau, Ministry of Labour, Japan

Year:

1996

Bibliographic source:

Japan Chemical Industry Ecology-Toxicology & Information Center, Japan (JETOC), January 1996

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

JAPAN: Guidelines for Screening Mutagenicity Testing Of Chemicals

Version / remarks:

comparable to OECD 471

Principles of method if other than guideline:

The mutagenicity assay was carried out according to the guidelines of Ministry of Labour, Japan (1979, 1985 and 1988) and performed by the following methods of Ames et al. (1975), Maron and Ames (1983) and Matsushima et al. (1980).

GLP compliance:

not specified

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

Adipic acid

EC Number:

204-673-3

EC Name:

Adipic acid

Cas Number:

124-04-9

Molecular formula:

C6H10O4

IUPAC Name:

adipic acid

Details on test material:

Test substance: 99% pure

Specific details on test material used for the study:

Miti No.: (2)-858
purity: guaranteed reagent
source: Tokyo Kasei Kogyo Co. LTD, Tokyo, Japan
lot No.: AL01

Method

Target gene:

histidine gene for Salmonella strains and tryptophane gene for E. coli strain

Species / strain

Species / strain 1

Species / strain / cell type:

S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and E. coli WP2

Species / strain 2

Species / strain / cell type:

S. typhimurium TA 1538

Metabolic activation:

with and without

Metabolic activation system:

Type and composition of metabolic activation system:
- source of S9 : male Sprague Dawley rats (weight about 200 g, seven weeks old) pretreated with sodium phenobarbital (3 i.p. injections at 30 or 60 mg/kg) and 5.6-benofravone (one i.p. injection at 80 mg/kg) before sacrifice. The S9 mix contained 4mM NADPH, 5mM G-6-P, 8 mM MgCl2, 33 mM KCl, 100 mM sodium phosphate buffer and 10% S9. The S9 mix was freshly prepared before the test.

Test concentrations with justification for top dose:

100, 200, 500, 1000, 2000, 5000, and 10000 µg/plate

Vehicle / solvent:

DMSO

Controls

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

Positive controls:

yes

Positive control substance:

4-nitroquinoline-N-oxide

9-aminoacridine

sodium azide

other: 2-(2-furyl)-3-(5-nitro-2-furyl)acrylamide and bleomycin for TA100, and TA102 without S9 mix; 2-aminoanthracene (0.5 to 10 µg/plate for all strains with S9)

Details on test system and experimental conditions:

The preincubation procedure in the Salmonella/mammalian microsome test was performed as described by Matsushima et al., 1980. The test compound dissolved in 0.05 or 1 ml of solvent was supplemented with 0.5 ml of S9 mix or 0.1 M phosphate buffer and 0.1 ml of the tester strains. developed in 1975 by Ames et al.
The mixtures of S9 and test chemcials were incubated at 37°C for 20 minutes, and then added to 2 mL of molten top agar (45°C). The contents of each tube were mixed and poured onto a minimal glucose agar plate immediately. The plates were incubated at 37°C for 48 hours, and then the number of revertant colonies on each plate was scored with an automated colony counter.

Evaluation criteria:

Two-hold rule criteria was used for data evaluation (Ames et al., 1975). The chemicals are ocnsidered to be mutagenic when a dose-related increase in revertant colony count is observed and the number of revertant colonies per plate with the test substance is more than twice that of the negative control (solvent control) and when a reproducibility of the test result is observed.
Mutagenic potency was calculated by the following equation and maximum value of mutagenic potency was expressed as a specific activity on the data sheet:
Mutagenic potency (induced revertants/mg test substance) = (number of induced revertants on the dose X - number of revertant on the solvent contro) / mg of test chemical on the dose X.

Results and discussion

Test results

Test results 1

Species / strain:

S. typhimurium TA 1535

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

at 10000 µg/plate

Vehicle controls validity:

valid

Positive controls validity:

valid

Test results 2

Species / strain:

S. typhimurium TA 1537

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

at 10000 µg/plate

Vehicle controls validity:

valid

Positive controls validity:

valid

Test results 3

Species / strain:

S. typhimurium TA 98

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity, but tested up to precipitating concentrations

Vehicle controls validity:

valid

Positive controls validity:

valid

Test results 4

Species / strain:

S. typhimurium TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

at 10000 µg/plate

Vehicle controls validity:

valid

Positive controls validity:

valid

Test results 5

Species / strain:

S. typhimurium TA 1538

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

at 10000 µg/plate

Vehicle controls validity:

valid

Positive controls validity:

valid

Test results 6

Species / strain:

E. coli WP2 uvr A

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity, but tested up to precipitating concentrations

Vehicle controls validity:

valid

Positive controls validity:

valid

Any other information on results incl. tables

Adipic acid gave no evidence of mutagenicity in any of the
bacterial strains used. Positive controls gave the expected results.

Applicant's summary and conclusion

Executive summary:

Adipic acid was not mutagenic in an Ames test performed by the Ministry of Labour (Japan, 1996) similar to OECD TG 471 in bacteria such as Salmonella typhimurium strains TA98, TA100, TA1535, TA1537 and TA1538 and Escherichia coli WP2 up to concentrations of 10 mg/plate with or without metabolic activation. Solvent and positive controls were functional in all experiments.