Registration Dossier

Data platform availability banner - registered substances factsheets

Please be aware that this old REACH registration data factsheet is no longer maintained; it remains frozen as of 19th May 2023.

The new ECHA CHEM database has been released by ECHA, and it now contains all REACH registration data. There are more details on the transition of ECHA's published data to ECHA CHEM here.

Diss Factsheets

Administrative data

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

Adipic acid was negative in several gene mutation tests on Salmonella typhimurium and E. coli strains, in a chromosomal aberration test and in a gene mutation test in mammalian cells. There is no indication of a mutagenic potential of adipic acid.

Link to relevant study records

Referenceopen allclose all

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
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
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
Target gene:
histidine gene for Salmonella strains and tryptophane gene for E. coli strain
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and E. coli WP2
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
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.
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
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
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
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
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
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

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

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.

Endpoint:
in vitro gene mutation study in bacteria
Remarks:
Type of genotoxicity: gene mutation
Type of information:
experimental study
Adequacy of study:
supporting study
Study period:
1982
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: No GLP, short documentation, purity not specified, similar to TG471
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Principles of method if other than guideline:
The standard S. typhimurium plate-incorporation assay was performed. The S9 mix used as an in vitro metabolic activator system contained 10% Aroclor 1254-induced liver S9 from male  Sprague-Dawley rats. Each substance was tested in the presence and in the  absence of S9 mix. In addition the tryptophan requiring E. coli strain  WP2  was tested for reversion to tryptophan independance. This test was performed by the same procedure as the S. typhimurium assay except that agar was supplemented with Oxoid nutrient broth to provide a trace of tryptophan. All platings were performed in duplicates and all tests were repeated on a different day. Concurrent positive controls were run with each test. The results were considered valid only if the positive control compound  induced increase in mutant counts to at least twice  background. The  following positive control compounds were used in the absence of S9:  2-nitrofluorene (5 or 10 µg per plate) for S. typhimurium strains TA98  and TA1538; sodium azide (0.5 or 1 µg) for TA100 and TA1535;  9-aminoacridine (50 or 100 µg) for TA1537; and AF-2 (furylframide, 0.1  µg) or N-methyl-N'-nitro-N-nitrosoguanidine (ENNG) (10 µg) for E. coli.  2-Anthramine (1 to 10 µg) was the positive control compound requiring S9  metabolic activation used for all bacterial strains.
GLP compliance:
not specified
Type of assay:
bacterial reverse mutation assay
Target gene:
S. typhimurium TA 1535, TA 1537, TA 1538, TA 98, TA 100: histidine synthesis; Escherichia coli WP2 : tryptophan synthesis
Species / strain / cell type:
other: S. typhimurium TA 1535, TA 1537, TA 1538, TA 98, TA 100, and Escherichia coli WP2
Metabolic activation:
with and without
Metabolic activation system:
Aroclor 1254-stimulated Sprague-Dawley rat liver S9 mix
Test concentrations with justification for top dose:
0.033, 0.10, 0.33, 1.0, 3.3 and 10 mg/plate
A maximum concentration of 10 mg/plate was used unless solubility and/or toxicity constraints dictated a lower maximum dose level. The dose levels used were based on the results of preliminary tests performed with all substances with S. typhimurium strain TA100.
Vehicle / solvent:
as solvent sodium phosphate buffer was used; if a substance was insoluble in water DMSO was used as solvent
Negative solvent / vehicle controls:
yes
Positive controls:
yes
Positive control substance:
other: sodium nitrite, nitrofurantoin, 1-naphtylamine
Details on test system and experimental conditions:
Ames test
Species / strain:
other: S. typhimurium TA 1535, TA 1537, TA 1538, TA 98, TA 100
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity nor precipitates, but tested up to recommended limit concentrations
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Species / strain:
E. coli WP2
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity nor precipitates, but tested up to recommended limit concentrations
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid

Adipic acid gave no evidence of mutagenicity in any of the bacterial strains used. Negative and positive controls were functional.

Executive summary:

Adipic acid was neither mutagenic nor cytotoxic in a study similar to OECD TG 471 in bacteria such as Salmonella typhimurium strains TA98, TA100, TA1535, TA1537 and TA1538 or Escherichia coli WP2 up to concentrations of 10 mg/plate with or without metabolic activator S9 (Prival 1991).

Endpoint:
in vitro gene mutation study in bacteria
Remarks:
Type of genotoxicity: gene mutation
Type of information:
experimental study
Adequacy of study:
key study
Study period:
1985
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
study well documented, meets generally accepted scientific principles, acceptable for assessment
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Principles of method if other than guideline:
The S. typhimurium pre-incubation assay was performed. The S9 mix used as an in vitro metabolic activator system S9 from male Sprague-Dawley rats. Each substance was tested in the presence and in the absence of S9 mix. In addition the trytophan requiring E. coli strain WP2  was tested. This test was performed by the same procedure as the S. typhimurium assay except that tryptophan was added to the top agar. Positive controls: AF-2, ENNG, 9-aminoacridine(9AC),  4-nitroquinoline-1-oxide (4nQO), benzo(a)pyrene (BaP), 2-aminoanthracene  (2AA), and 2-nitrofluorene (12NF). All tests were performed in duplicates.
GLP compliance:
not specified
Type of assay:
bacterial reverse mutation assay
Specific details on test material used for the study:
purity: 99%
source: Tokyo Kasei Kogyo Co. LTD, Tokyo, Japan
Target gene:
histidine gene
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and E. coli WP2
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 220 g, seven weeks old) pretreated with polychlorinated biphenyl (KC 500) at a dose of 500 mg/kg bw 5 days before sacrifice. The S9 mix contained 0.1 mL of S9/mL.
0.5 mL of S9 mix or sodium phosphate buffer (pH 7.4) were added to a sterile test tube containing 0.1 mL of various concentrations of the chemicals.
Test concentrations with justification for top dose:
1, 5, 10, 50, 100, 500, 1000 and 5000 µg/plate
Vehicle / solvent:
DMSO
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
Positive controls:
yes
Positive control substance:
4-nitroquinoline-N-oxide
9-aminoacridine
N-ethyl-N-nitro-N-nitrosoguanidine
benzo(a)pyrene
other: 2-(2-furyl)-3-(5-nitro-2-furyl)acrylamide (AF-2; 0.01 µg/plate for TA100 without S9 and 0.05 µg/plate for WP2 and TA98 without S9); 2-aminoanthracene (2-AA; 5.0 µg/plate TA1535 and WP2 with S9)
Details on test system and experimental conditions:
Salmonella/mammalian microsome test developed in 1975 by Ames et al.
The mixtures of S9 and test chemcials were incubated in a shaker water bath 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.
Species / strain:
S. typhimurium TA 1535
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity nor precipitates, but tested up to recommended limit concentrations
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Species / strain:
S. typhimurium TA 1537
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity nor precipitates, but tested up to recommended limit concentrations
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Species / strain:
S. typhimurium TA 98
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity nor precipitates, but tested up to recommended limit concentrations
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Species / strain:
S. typhimurium TA 100
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity nor precipitates, but tested up to recommended limit concentrations
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Species / strain:
S. typhimurium TA 1538
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity nor precipitates, but tested up to recommended limit concentrations
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Species / strain:
E. coli WP2 uvr A
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity nor precipitates, but tested up to recommended limit concentrations
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid

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

Executive summary:

Adipic acid was not mutagenic in a study similar to OECD TG 471 in bacteria such as Salmonella typhimurium strains TA98, TA100, TA1535, TA1537 and TA1538 or Escherichia coli WP2 up to concentrations of 5 mg/plate with or without metabolic activator S9. Negative and positive controls were functional in all experiments (Shimizu et al. 1985).

Endpoint:
in vitro cytogenicity / chromosome aberration study in mammalian cells
Remarks:
Type of genotoxicity: chromosome aberration
Type of information:
experimental study
Adequacy of study:
key study
Study period:
1974
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: No GLP, but good documentation, purity not specified, no metabolic activation
Principles of method if other than guideline:
Human embryonic lung fibroblast cultures (WI-38) were suspended in tissue culture medium and plated. The test compound was added at three dose levels using three bottles for each level, 24 hours after plating. A preliminary determination of tissue culture toxicity was performed (cytotoxic effects were observed at 400 mg/l). Cells were incubated at 37 degree Celsius and examined twice daily to determine when an adequate number of mitoses were present. Cells were harvested and fixed (3:1 absolute methanol : glacial acetic acid). The specimens were centrifuged, decanted, and suspended in acetic acid-orcein stain and dropped on a slide. The preparations were examined by microscopy. Cells in anaphase were observed for non-disjunction as indicative of cytogenetic damage. Analyzed aberrations include bridges, pseudochiasmata, multipolar cells, and acentric fragments. The positive control was triethylene melamine (TEM) and the negative control was saline. 100 cells were investigated per dose.
GLP compliance:
no
Type of assay:
in vitro mammalian chromosome aberration test
Species / strain / cell type:
other: human fibroblasts
Details on mammalian cell type (if applicable):
human fibroblasts (WI-38)
Metabolic activation:
without
Test concentrations with justification for top dose:
0, 2, 20, 200 mg/l
Vehicle / solvent:
0.85% saline
Negative solvent / vehicle controls:
yes
Positive controls:
yes
Positive control substance:
triethylenemelamine
Species / strain:
other: human fibroblasts
Metabolic activation:
without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
other: 400 mg/l
Vehicle controls validity:
valid
Positive controls validity:
valid
Remarks on result:
other: other: human fibroblasts (WI-38)
Remarks:
Migrated from field 'Test system'.

Negative and positive controls were functional. The negative controls contained two cells with bridges one of which contained an acentric fragment. The test compound was negative except for one cell which contained a bridge at the high  

dose level. In summary, the compound produced no significant aberration.

 conc. (mg/L)  mitotic index*  no. of cells  % cells with acentric frag.  % cells with bridges  % multipolar cells  % cells other aberrations  % total cells with aberrations**
 vehicle saline  1  100  1  2  0  0  3
 2.0  1  100  0  0  0  0  0
 20.0  1  100  0  0  0  0  0
 200.0  1  100  0  1  0  0  0
 positive control TEM 0.1  1  100  3  12  0  0

 15

* percent of cells in mitosis: 200 cells observed per concentration

** duplicate aberrations in a single cell will cause this to be a % less than a summation of the % aberrations seen

Executive summary:

Adipic acid was inactive in a cytogenetic assay using human embryonic lung fibroblast cells (WI-38) and compound concentrations up to 200 mg/l. Cytotoxicity was observed at 400 mg/l. No metabolic activation system was used in these experiments; the positive and negative controls were functional (Litton Bionetics, Inc. 1974).

Endpoint:
in vitro gene mutation study in mammalian cells
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 476 (In Vitro Mammalian Cell Gene Mutation Test)
Deviations:
no
Qualifier:
according to guideline
Guideline:
EU Method B.17 (Mutagenicity - In Vitro Mammalian Cell Gene Mutation Test)
Qualifier:
according to guideline
Guideline:
EPA OPPTS 870.5300 - In vitro Mammalian Cell Gene Mutation Test
GLP compliance:
yes (incl. QA statement)
Type of assay:
mammalian cell gene mutation assay
Specific details on test material used for the study:
content: 99.92%
Target gene:
HPRT
Species / strain / cell type:
Chinese hamster lung fibroblasts (V79)
Additional strain / cell type characteristics:
not applicable
Metabolic activation:
with and without
Metabolic activation system:
The S9 liver microsomal fraction was prepared at BSL BIOSERVICE GmbH. Male Wistar rats were induced with Phenobarbital (80 mg/kg bw) and B-Naphthoflavone (100 mg/kg bw) for three consecutive days by oral route.
Test concentrations with justification for top dose:
The selection of the concentrations was based on data from the pre-experiment. In all experiments 10 mM was selected as the highest concentration.
Experiment 1:
0.1, 0.25, 0.5, 1, 2.5, 5, 7.5 and 10 mM with and without metabolic activation
Experiment 2:
0.5, 0.75, 1, 2,5, 4, 7, 8.5 and 10 mM with metabolic activation
0.1, 0.25, 0.5, 1, 2.5, 5, 7.5 and 10 mM without metabolic activation
According to OECD Guidelines at least 8 concentrations ofthe test item were set up in the experiments with and without metabolic activation.
Vehicle / solvent:
The test item was dissolved in cell culture medium (MEM + 0% FBS 4h treatment; MEM + 10% FBS 20h treatment) and diluted prior to treatment. The solvent was compatible with the survival of the cells and the S9 activity.
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
Positive controls:
yes
Positive control substance:
ethylmethanesulphonate
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
Positive controls:
yes
Positive control substance:
7,12-dimethylbenzanthracene
Details on test system and experimental conditions:
Pretest for Toxicity:
The toxicity of the test item was detennined in a pre-experiment. Eight concentrations [0.1, 0.25, 0.5, 1.0,2.5,5.0,7.5, 10 mM] were tested without metabolic activation. The experimental conditions in this pre-experiment were the same as described below for the main experiment.
Evaluation criteria:
A test is considered to be negative if there is no biological relevant increase in the number of mutants.
There are several criteria for determining a positive result:
a reproducible three times higher mutation frequency than the solvent control for at least one of the concentrations; a concentration related increase of the mutation frequency; such an evaluation may be considered also in the case that a three-fold increase of the mutant frequency is not observed;
According to the OECD guidelines, the biological relevance of the results is the criterion for the interpretation of results, a statistical evaluation of the results is not regarded as necessary.
Species / strain:
Chinese hamster lung fibroblasts (V79)
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity nor precipitates, but tested up to recommended limit concentrations
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Additional information on results:
No precipitation of the test item was noted in the experiments.
Remarks on result:
other: strain/cell type: Chinese hamster lung fibroblasts (V79)
Remarks:
Migrated from field 'Test system'.

Toxicity:

In experiment I a growth inhibition (reduction of relative growth to 61.0%) was observed after the treatment with the test item at a concentration of 0.25 mM (without metabolic activation). Due to the overall consistency of the toxicity data at lower and at higher concentrations this finding was considered as not biologically relevant. No biologically relevant growth inhibition was observed after the treatment with the test item in experiment I with metabolic activation and in experiment II with and without metabolic activation.

Mutagenicity:

DMBA (1.0 Ilg/mL (experiment I) and 2.0 Ilg/mL (experiment II)) and EMS (300 Ilg/mL) were used as positive controls and showed distinct and biologically relevant effects in mutation frequency.

In conclusion, in the described in vitro cell gene mutagemcIty test under the experimental conditions reported, the test item Adipic Acid is considered to be nonmutagenic in the HPRT locus using V79 cells of the Chinese Hamster.

 dose group  mutation factor exp.1 (without S9 mix)  mutation factor exp. 1 (with S9 mix)  mutation factor exp. 2 (without S9 mix)  mutation factor exp. 2 (with S9 mix)
 control  0  0  0  0
 0.10  2.06  0.61  1.39  -
 0.25  1.40  1.52  1.63  -
 0.5  1.75  1.00  1.09  0.50
 0.75  -  -  -  0.57
 1.0  2.06  0.91  1.31  0.55
 2.5  1.31  1.13  1.55  0.53
 5.0  1.44  1.10  1.35  0.75
 7.0  -  -  -  0.77
 7.5  0.97  1.58  1.18  -
 8.5  -  -  -  0.34
 10.0  1.28  1.22  1.12  0.43
 EMS 300 µg/mL  13.47  -  87.03  -
 DMBA 1 µg/mL  -  8.50  -  -
 DMBA 2 µg/mL  -  -  -  30.56
Executive summary:

Adipic acid was investigated in an OECD TG 476 study in Chinese hamster V79 cells in the absence and in the presence of metabolic activation (S9) in concentrations of up to and including 10 mM. No precipitation of the test item was noted in any experiment; no biological relevant growth inhibition was observed with and without metabolic activation. In both experiments no biologically relevant increase of mutants was found after treatment with the test item. DMBA and EMS were used as positive controls and showed distinct and biologically relevant effects in mutation frequencies.

In conclusion, adipic acid is considered to be non-mutagenic in the HPRT locus using V79 cells.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (negative)

Genetic toxicity in vivo

Description of key information

Adipic acid was not mutagenic in in vivo cytogenetic studies where groups of five rats were dosed with adipic acid doses up to 5000 mg/kg bw (acute studies) and with doses up to 2500 mg/kg bw/day (five-days subacute studies).

Link to relevant study records
Reference
Endpoint:
in vivo mammalian somatic cell study: cytogenicity / bone marrow chromosome aberration
Type of information:
experimental study
Adequacy of study:
key study
Study period:
1974
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: No GLP but overall good documentation, purity not specified, no positive control for every experiment.
Principles of method if other than guideline:
Groups of  5 treated and 3 control animals were used. Animals were killed  6, 24 and 48 hours after a single administration in the acute study. In  the subacute study 5 doses, 24 hours apart, were administered and animals  were killed 6 hours after the last dose. 
GLP compliance:
no
Type of assay:
mammalian bone marrow chromosome aberration test
Specific details on test material used for the study:
compound FDA 71-50, Adipic acid, food processing quality, as supplied by the Food and Drug Administration
Species:
rat
Strain:
not specified
Sex:
male
Route of administration:
oral: gavage
Duration of treatment / exposure:
Acute study: single dosing; subacute study: once a day for 5 consecutive days
Frequency of treatment:
Acute study: single dosing; subacute study: once a day for 5 consecutive days
Post exposure period:
Animals were killed  6, 24 and 48 hours after a single administration in the acute study. In  the subacute study 5 doses, 24 hours apart, were administered and animals  were killed 6 hours after the last dose. 
Remarks:
Doses / Concentrations:
Test 1: acute and subacute: 3.75, 37.5, 375 mg/kg bw/day; Test 2: acute 5000 mg/kg bw and subacute 2500 mg/kg bw/day
Basis:

No. of animals per sex per dose:
Groups of  5 treated and 3 control animals were used.
Control animals:
yes, concurrent vehicle
Positive control(s):
positive control animals were treated with 0.3 mg/kg bw TEM (triethylenemelamine) by intraperitoneally incection
Tissues and cell types examined:
bone marrow
Details of tissue and slide preparation:
Four hours after the last  compound administration, and two hours prior to killing, each animal was  given 4 mg/kg bw of colcemid intraperitoneally in order to arrest the  bone marrow cells in C-mitosis. The marrow "plug" was removed and  aspirated into Hanks' balanced salt solution. The specimen were  centrifuged and resuspended in hypotonic 0.5% KCl. The specimens were  placed in a 37 degree Celsius water bath in order to swell the cells.  Following centrifugation the cells were resuspended in a fixative (3:1  absolute methanol : glacial acetic acid) and again centrifuged. Cells were  resuspended and placed at 4 degree Celsius overnight. The following day  cells were again centrifuged and freshly prepared fixative was added. The  suspension was dropped onto a slide and ignited by an alcohol burner and  allowed to flame. Slides were stained with 5% Giemsa solution. The  preparations were examined by microscopy. The chromosomes of each cell  were counted and only diploid cells were analyzed. They were scored for  chromatid gaps and breaks, chromosome gaps and breaks, reunions, cells  with greater than ten aberrations, polyploidy, pulverization, and other  chromosomal aberrations which were observed. Fifty metaphase spreads were  scored per animal. Mitotic indices were obtained by counting at least 500  cells and the ratio of the number of cells in mitosis / the number of  cells observed was expressed as the mitotic index. Negative and positive  (TEM) controls were run in each experiment. Two tests were performed at different time intervals.
Sex:
male
Genotoxicity:
negative
Toxicity:
not specified
Vehicle controls validity:
valid
Negative controls validity:
not applicable
Positive controls validity:
valid
Remarks:
Triethyl Melamine (TEM)

Table 1: Acute study I with single gavage dosing (3.75, 37.5 and 375 mg/kg bw):

The negative control group cells contained no aberrations. Adipic acid produced no aberrations except for one cell containing a break in the 6-hour sample of  the intermediate dose level. The expected severe chromosomal damage was observed for the positive control group (triethylene melamine treated animals). The mitotic indices  

were within normal limits. Negative and positive controls were functional.

 dosage (mg/kg bw)  time*  no. of animals no. of cells per animal  mitotic index**  % cells with breaks  % cells with other aberrations  % total cells with aberrations
 negative control saline  6/24/48  3/3/3  150  6/4/8  0/0/0  0/0/0  0/0/0
 3.75  6/24/48  5/5/5  250  7/4/6  0/0/0  0/0/0  0/0/0
 37.5  6/24/48  5/5/5  250  5/7/6  0.4/0/0  0/0/0  0/0/0
 375  6/24/48  5/5/5  250  5/6/4  0/0/0  0/0/0  0/0/0

 positive control

TEM 0.3

 48  5  250  4  0  22

 45

* time of kill after single gavage application (hours)

**percent cells in mitosis: 500 cells observed/animal

Table 2: Subacute study I (5 days) with 5 gavage dosings (3.75, 37.5 and 375 mg7kg bw/day): 

The negative control group and the low level test group contained no aberration. The intermediate level contained one cell with a reunion and one cell that  

was polyploid. The highest level contained three cells with breaks and one fragment. These were considered to be within the normal limits of the  

historical negative controls of the laboratory. Negative control was functional, no positive control.  

 dosage (mg/kg bw)#  no. of animals no. of cells per animal  mitotic index**  % cells with breaks  % cells with reunion  % cells with other aberrations % total cells with aberrations 
 negative control saline 3  150  6  0  0  0  0
 3.75 5  250  5  0  0  0  0
 37.5 5  250  5  0  0.4  0.4 (P)  0.8
 375 5  250  4  1.2  0  0.4 (f)  1.6

# dosage as gavage application 1x/day for 5 days

P - cells that have polyploidy

f - cells that have fragmentaion

** percent cells in mitosis: 500 cells observed/animal

Table 3: Acute study II: Adipic acid was administered at a single dose of 5000 mg/kg bw. The compound produced no aberrations except for 3 cells with polyploidy (2 in the 6-hour sample and 1 in the 24-hour). Neither the variety nor the number of these aberrations differed significantly from the negative controls (polyploidy observed in 4 cells). Negative and positive controls were functional.

 dosage (mg/kg bw)  time*  no. of animals no. of cells per animal  mitotic index**  % cells with breaks  % cells with other aberrations  % total cells with aberrations
 negative control saline  6/24/48  3/3/3  150  7.47/4.5/4.5  0/0/0  1pp/2pp/1pp  1 (0.66)/2 (1.33)/1 (0.66)
 5000  6/24/48  5/5/5  250  5.51/4.03/4.13  0/0/0  2pp/1pp/0  2 (0.8)/1 (0.4)/0

 positive control

TEM 0.3

24  5  250  1.62  4 (1.6)  >27 (10.8), 9f, 1pp

 86 (34.4)

P - cells that have polyploidy

f - cells that have fragmentaion

** percent cells in mitosis: at least 500 cells observed/animal

Table 4: Subacute study (5 days, 2500 mg/kg bw/day). Only 218 metaphases have been evaluated. The compound produced no aberrations except for 1 cell with polyploidy. Polyploidy was also observed in the negative control group. These are considered to be within the normal limits of the historical negative controls. Negative control was functional, no  positive 

control.

 dosage (mg/kg bw)#  no. of animals no. of cells per animal  mitotic index**  % cells with breaks  % cells with reunion  % cells with other aberrations % total cells with aberrations 
 negative control saline 3  150  5.33  0  0  1 (pp) (0.66)  1 (0.66)
 2500 5  218  2.98  0  0  1 (pp) (0.46)  1 (0.46)

P - cells that have polyploidy

f - cells that have fragmentaion

** percent cells in mitosis: 500 cells observed/animal

In summary, adipic acid can be considered non-mutagenic as measured by the cytogenetic test.

Executive summary:

Adipic acid was not mutagenic in in vivo cytogenetic studies where groups of five male rats were gavaged with adipic acid doses up to 5000 mg/kg bw (acute studies) and with doses up to 2500 mg/kg bw/day (five-days subacute studies). 200 to 500 metaphase chromosomes of bone marrow cells per dose were scored for chromatid gaps and breaks, chromosome gaps and breaks, reunions, cells with greater than ten aberrations, polyploidy, pulverization and other chromosomal aberrations. The mitotic indices for all dose groups were considered to be within the normal limits of the controls and there was no evidence of chromosomal damage. The positive control groups, performed only during the acute studies, were functional (Litton Bionetics, Inc. 1974).

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (negative)

Additional information

In vitro:

Hazards identified by OECD/ICCA high production volume chemicals program in 2004:

"Adipic acid was neither mutagenic nor cytotoxic in studies similar to OECD TG 471 in bacteria such as Salmonella typhimurium strains TA98, TA100, TA1535, TA1537 and TA1538 or Escherichia coli WP2 up to concentrations of 10 mg/plate with or without metabolic activator S9. Negative and positive controls were functional in all experiments (Mortelmans and Griffin 1982; Prival et al. 1991 ; Shimizu et al. 1985).

Adipic acid was negative in a yeast gene mutation assay using Saccharomyces cerevisiae D3 as a reporter strain in the absence of S9-mix and adipic acid concentrations up to 200 mg/l. Cytotoxicity was not mentioned. The positive and negative controls were functional (Litton Bionetics, Inc. 1974).

Adipic acid was also inactive in a cytogenetic assay using human embryonic lung fibroblast cells (WI-38) and compound concentrations up to 200 mg/l. Cytotoxicity was observed at 400 mg/l. No metabolic activation system was used in these experiments and the positive and negative controls were functional (Litton Bionetics, Inc. 1974)."

Updated relevant information:

Negative results in well performed bacterial gene mutation tests (Ames tests) were also found by e.g. NTP (2015) and the Ministry of Labour, Japan (1996).

Adipic acid was investigated in an OECD TG 476 study in Chinese hamster V79 cells in the absence and in the presence of metabolic activator (S9) up to concentrations of 10 mM. No precipitation of the test item was noted in any experiment; no biological relevant growth inhibition was observed with and without metabolic activation. In both experiments no biologically relevant increase of mutants was found after treatment with the test item. DMBA and EMS were used as positive controls and showed distinct and biologically relevant effects in mutation frequencies. In conclusion, adipic acid is considered to be non-mutagenic in the HPRT locus using V79 cells (BSL Bioservice, 2009).

In vivo:

Hazards identified by OECD/ICCA high production volume chemicals program in 2004:"Adipic acid was investigated in a host mediated assay with Salmonella typhimurium TA-1530 and G-46 or Saccharomyces cerevisiae D3 as indicator strains. In an acute and subacute study groups of 10 male mice were dosed with 3.75, 37.5 and 375 mg/kg bw/day for one and 5 days, respectively. Adipic acid produced no significant increase in mutation frequencies in any experiment, except when using Saccharomyces cerevisiae D3 in the acute study. In this case an increased frequency of mutations as well as dose response was observed. In further experiments in the same study animals were dosed with 5000 mg/kg bw once and with 2500 mg/kg bw/day for 5 days, respectively. In these studies the results were negative for all three indicator strains TA-1530, G-46 and Saccharomyces cerevisiae D3 in both, the acute and subacute, experiments. The positive control groups, employed only during the acute studies, were functional (Litton Bionetics, Inc. 1974).

Adipic acid was not mutagenic in in vivo cytogenetic studies where groups of five rats were dosed with adipic acid doses up to 5000 mg/kg bw (acute studies) and with doses up to 2500 mg/kg bw/day (five-days subacute studies). 200 to 500 metaphase chromosomes per dose were scored for chromatid gaps and breaks, chromosome gaps and breaks, reunions, cells with greater than ten aberrations, polyploidy, pulverization, and other chromosomal aberrations. The mitotic indices for all dose groups were considered to be within the normal limits of the controls and there was no evidence of chromosomal damage. The positive control groups, performed only during the acute studies, were functional (Litton Bionetics, Inc. 1974).

Adipic acid was administered to groups of 10 male rats in a dominant lethal assay. Each treated male rat was mated with two virgin female rats each week for seven (subacute study) or eight (acute study) weeks. Two weeks after mating, female rats were sacrificed and the fertility index, preimplantation loss and lethal effects on the embryos were determined and compared with those same parameters calculated from control animals. In an acute study (3.75, 37.5 and 375 mg/kg bw) a decrease in average implantations at week 1 and 4, and corpora lutea at week 4 and 7 were seen only in the intermediate dose level. Increase in preimplantation losses were shown at week 1 for both the low and intermediate dose groups with no changes at any other week and parameter. In a five days subacute study with the same doses significant differences between the negative control and experimental groups were shown in a few instances, but no clear indications of change were seen. In a second test (acute single dose of 5000 mg/kg bw and subacute five doses of 2500 mg/kg bw/day) the values from those animals dosed with adipic acid did not significantly vary from those obtained from the negative control. Positive control groups, performed during the acute studies, gave the expected results. In summary, adipic acid does not induce dominant lethal mutations in doses up to 5000 mg/kg bw (Litton Bionetics, Inc. 1974)."

Updated relevant information:

None

Justification for classification or non-classification

Based on negative results in vitro for gene mutations in bacteria and mammalian cells and chromosomal aberrations in mamalian cells, as well as in vivo experiments with acute and subacute exposure to adipic acid, no classification is required.