Carboxylic acids, di-, C4-6

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

Study period:

1986

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1986

Report date:

1986

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

not specified

Principles of method if other than guideline:

Bibliography: Ames et al., Mut.Research. 31: 347-364, 1975
Appropriate positive controls included

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

Carboxylic acids, di-, C4-6

EC Number:

271-678-5

EC Name:

Carboxylic acids, di-, C4-6

Cas Number:

68603-87-2

Molecular formula:

C5H8O4, C4H6O4, C6H10O4

IUPAC Name:

Carboxylic acids, C4-6 di-

Details on test material:

Sample idetification: AGS Mixture:
Adipic acid: 6-9%
Glutaic acid 30-35%
Succinic acid 8-11%
Nitric acid 0.5 - 3%
Lot: FIT-85-45,46,47; white solid powder

Specific details on test material used for the study:

test item: AGS-Mixture (Adipic acid: 6-9%; Glutaric acid 30-35%; Succinic acid 8-11%; Nitric acid 0.5 - 3%) used as 50% aqueous solution

Method

Target gene:

Histidine auxotrothy

Species / strain

Species / strain 1

Species / strain / cell type:

S. typhimurium TA 1535, TA 1537, TA 98 and TA 100

Species / strain 2

Species / strain / cell type:

S. typhimurium TA 1538

Metabolic activation:

with and without

Metabolic activation system:

rat liver S9 mix obtained from Aroclor 1254-treated Sprague Dawley rats

Test concentrations with justification for top dose:

30, 100, 300, 1000, 3000 µg/plate
These dose levels were determined in a preliminary toxicity screen with 10, 167, 500, 1667 and 5000 µg/plate.

Vehicle / solvent:

water

Controls

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

Remarks:

Water

Positive controls:

yes

Positive control substance:

other: Sodium acide, 9-aminoacridine, 2-nitrofluorene, 2-anthramine

Details on test system and experimental conditions:

plate incorporation assay

Evaluation criteria:

A positive result is defined as a reproducible, dose-related increase in the number of histidine-independent colonies with at least one dose point inducing a mutant frequency value that is two-fold the solvent control value.

Statistics:

Significance at the 95% conficence limits is determined by the program developed by Moore and Felton (1983).

Results and discussion

Test results

Species / strain:

other: all strains tested

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

other: all strains/cell types tested

Any other information on results incl. tables

Bacterial toxicity was investigated in a pilot experiment; no relevant colonies at 5000 µg/plate; high dose for main experiment: 3000µg/plate. All solvent and positive controls showed the expected results.

The results of the test article, AGS Mixture, were negative in strains TA 1535, 1537, 98 and 100 of S. typhimurium with and without metabolic activation at all doses tested (highest dose: 3000 µg/plate).

In the initial assay, strain 1538 exhibited an increase in mutant frequency at the hghest dose (2.5 fold above the concommitant control) without metaboilic activation. Two subsequent retests failed to confirm the initial observation (see Table 1)

Overall the authors concluded that the test compound is not mutagenic in the Ames test.

Table 1: summary of results

dose level (µg/plate)	S9 -mix ( + or -)	TA1535	TA1537	TA98	TA100	TA1538 (1.exp)	TA1538 (2. exp)	TA1538 (3.exp)
water	-	14	13	28	141	14	12	12
water	+	9	11	44	159	20	-	-
sodium acide, 10	-	1107	-	-	1152	-	-	-
9 -Aminoacridine, 150	-	-	1496	-	-	-	-	-
2 -Nitrofluorene, 5	-	-	-	486	-	507	446	551
2 -Anthramine, 2.5	+	250	232	1544	1467	1436	-	-
AGS mix, 30	-	17	12	26	150	13	6	12
AGS mix, 100	-	13	10	24	133	15	8	12
AGS mix, 300	-	16	12	24	144	15	5	9
AGS mix, 1000	-	14	15	32	148	15	11	16
AGS mix, 3000	-	17	14	42	140	35	21	11
AGS mix, 30	+	10	10	38	119	18	-	-
AGS mix, 100	+	11	10	42	188	25	-	-
AGS mix, 300	+	8	13	34	190	13	-	-
AGS mix, 1000	+	8	18	43	181	31	-	-
AGS mix, 3000	+	11	14	42	197	30	-	-

AGS mixture was tested as 50% aqueous solution.

Applicant's summary and conclusion

Executive summary:

Mutagenic activity of dicarboxylic acid mixture (AGS-Mixture, 50%) was investigated in a study similar to OECD TG 471 in bacteria (Salmonella typhimurium strains TA98, TA100, TA1535, TA1537 and TA1538) with or without metabolic activator S9. Cytotoxicity was observed at 5000 µg/plate. Negative and positive controls were functional in all experiments (Barfknecht 1986, BASF 1990).

In the initial assay, strain TA1538 exhibited a dose related increase in mutant frequency that reached 2.5-fold the solvent control background value without metabolic activation. However, two subsequent retests failed to confirm the initial observation. All other tests were negative. The authors concluded that dicarboxylic acid mixture is not mutagenic in the 5 strains tested with and without metabolic activation at dose levels of 30, 100, 300, 1000 and 3000 µg/plate.