Aminocaproic acid

Endpoint summary

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

The test substance was considered to be non-mutagenic with and without metabolic activation in bacteria (reference 7.6.1 -1).

Link to relevant study records

Reference

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

06 October 2016 - 08 November 2016

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Version / remarks:

1997

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)

Version / remarks:

Council Regulation (EC) No 440/2008

Deviations:

no

Qualifier:

according to guideline

Guideline:

JAPAN: Guidelines for Screening Mutagenicity Testing Of Chemicals

Version / remarks:

cf. Sofuni, 1993

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Type of assay:

bacterial reverse mutation assay

Species / strain / cell type:

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

Metabolic activation:

with and without

Metabolic activation system:

S9 mix induced by Aroclor 1254 in livers of rats

Test concentrations with justification for top dose:

5, 15.8, 50, 158, 500, 1580 and 5000 µg/plate
top dose: in order that initial treatments were performed up to the maximum recommended concentration according to current regulatory guidelines (OECD 471, 1997)

Vehicle / solvent:

- Vehicle/solvent used: ultrapure water
- Justification for choice of solvent/vehicle: based on the available information from the preliminary solubility test

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

4-nitroquinoline-N-oxide

9-aminoacridine

sodium azide

other: daunomycin, 2-aminoanthracene

Details on test system and experimental conditions:

METHOD OF APPLICATION: in agar (plate incorporation)

DURATION
- Exposure duration: 2 to 3 days

NUMBER OF REPLICATIONS: 3 (two independent experiments)

DETERMINATION OF CYTOTOXICITY
- Method: counting numbers of revertants

OTHER:
-S9 concentration: 1st series 10%, 2nd series 30%

Evaluation criteria:

A test material was to be defined as negative or non-mutagenic in this assay if
- The assay was to be considered valid, and
- "no" or "weak increases" occurred in the test series performed ("weak increases" randomly occur due to experimental variation)

For valid data, the test material was considered to be positive or mutagenic if:
- a dose dependent (over at least two test material concentrations) increase in the number of revertants was induced, the maximal effect was a "clear increase", and the effects were reproduced at similar concentration levels in the same test system, or
- "clear increases" occurred at least at one test material concentration, higher concentrations showed strong precipitation or cytotoxicity, and the effects were reproduced at the same concentration level in the same test system.

Key result

Species / strain:

S. typhimurium TA 1535

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

Positive controls validity:

valid

Key result

Species / strain:

S. typhimurium TA 1537

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

Positive controls validity:

valid

Key result

Species / strain:

S. typhimurium TA 98

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

Positive controls validity:

valid

Key result

Species / strain:

S. typhimurium TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

Positive controls validity:

valid

Key result

Species / strain:

E. coli WP2 uvr A

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
- Precipitation: No test material precipitate was observed on the plates at any of the doses tested in either the presence or absence of S9-mix.

Table 1:  Revertants per plate (Mean ± SD) - 1^st series

Metabolic Activation	Test Material	Concentr. [µg/plate]	Revertants per plate (Mean ± SD)
			TA 98	TA 100	TA 1535	TA 1537	WP2 uvrA
Without Activation	H2O		41± 3	113 ± 13	29 ± 6	23 ± 2	34 ± 8
	Art A2504	5.00	35 ± 5	115 ± 5	26 ± 2	28 ± 4	31 ± 5
		15.8	33 ± 3	107 ± 15	37 ± 2	27 ± 4	35 ± 4
		50.0	44 ± 1	121 ± 17	36 ± 7	27 ± 9	39 ± 7
		158	39 ± 7	114 ± 10	32 ± 5	28 ± 8	28 ± 1
		500	35 ± 7	108 ± 5	29 ± 2	28 ± 6	34 ± 8
		1580	36 ± 7	115 ± 7	31 ± 7	23 ± 2	29 ± 4
		5000	41 ± 7	109 ± 19	26 ± 2	27 ± 2	35 ± 7
	DAUN	1.00	464 ± 94
	NaN3	2.00		1609 ± 103	949 ± 96
	9-AA	50.0				1926 ± 339
	NQO	2.00					2221 ± 218
With Activation	H2O		50 ± 9	125 ± 12	29 ± 5	27 ± 4	33 ± 5
	Art. A2504	5.00	46 ± 4	134 ± 23	29 ± 6	25 ± 6	37 ± 7
		15.8	43 ± 8	127 ± 7	28 ± 1	32 ± 9	32 ± 5
		50.0	30 ± 1	143 ± 5	30 ± 4	28 ± 3	37 ± 11
		158	48 ± 7	118 ± 14	30 ± 2	30 ± 2	37 ± 1
		500	47 ± 3	125 ± 6	28 ± 4	23 ± 2	38 ± 1
		1580	48 ± 8	135 ± 14	32 ± 2	23 ± 1	37 ± 1
		5000	48 ± 8	135 ± 6	33 ± 1	28 ± 4	36 ± 1
	2-AA	2.00	301 ± 35	517 ± 62
	2-AA	5.00			170 ± 16	156 ± 5
	2-AA	10.0					298 ± 29

NaN3: Sodium azide, 2-AA: 2-Aminoanthracene, 9 -AA: 9-Aminoacridine, DAUN: Daunomycin, NQO: 4-NitroquinoIine-N-oxide

Table 2:  Revertants per plate (Mean ± SD) - 2^nd series

Metabolic Activation	Test Material	Concentr. [µg/plate]	Revertants per plate (Mean ± SD)
			TA 98	TA 100	TA 1535	TA 1537	WP2 uvrA
Without Activation	H2O		31± 7	115 ± 5	30 ± 4	25 ± 3	35 ± 6
	Art A2504	50.0	27 ± 8	98 ± 9	37 ± 4	25 ± 6	32 ± 4
		158	28 ± 4	103 ± 11	31 ± 7	26 ± 6	36 ± 4
		500	21 ± 7	113 ± 10	37 ± 5	25 ± 3	41 ± 9
		1580	27 ± 6	111 ± 14	27 ± 8	27 ± 8	35 ± 6
		5000	22 ± 8	115 ± 4	35 ± 3	25 ± 4	24 ± 4
	DAUN	1.00	301 ± 10
	NaN3	2.00		1591 ± 73	914 ± 92
	9-AA	50.0				1061 ± 196
	NQO	2.00					1800 ± 399
With Activation	H2O		30 ± 8	107 ± 7	24 ± 5	30 ± 2	46 ± 7
	Art. A2504	50.0	28 ± 4	129 ± 10	23 ± 7	26 ± 9	42 ± 12
		158	34 ± 9	118 ± 9	21 ± 7	29 ± 5	44 ± 7
		500	29 ± 10	100 ± 5	25 ± 1	31 ± 5	32 ± 5
		1580	28 ± 5	106 ± 7	26 ± 9	28 ± 7	41 ± 3
		5000	33 ± 3	113 ± 3	23 ± 4	26 ± 7	37 ± 7
	2-AA	2.00	164 ± 42
	2-AA	5.00		949 ± 108
	2-AA	10.0			136 ± 11	226 ± 28	217 ± 36

NaN3: Sodium azide,2-AA: 2-Aminoanthracene,9 -AA:9-Aminoacridine,DAUN:Daunomycin,NQO:4-NitroquinoIine-N-oxide

Table 3: Historical data - negative controls

Strain	TA 98		TA 100		TA 1535		TA 1537		WP2 uvrA
S9 Mix	Without	With	Without	With	Without	With	Without	With	Without	With
Compound	Solvent	Solvent	Solvent	Solvent	Solvent	Solvent	Solvent	Solvent	Solvent	Solvent
Total Plates	380	379	180	179	176	176	304	304	388	400
Number of Values	89	89	39	39	38	38	70	70	91	94
Minimum	28	31	23	19	15	13	23	26	89	95
Maximum	49	54	54	39	29	33	49	51	147	166
Mean	37	42	31	28	24	25	33	38	111	120
Standard Deviation	4.3	5.6	3.9	4.4	3.4	4.7	5.3	5.3	12.2	11.3

Table 4: Historical data - positive controls

Strain	TA 98		TA 100		TA 1535		TA 1537		WP2 uvrA
S9 Mix	Without	With	Without	With	Without	With	Without	With	Without	With
Compound	DAUN	2-AA	NaN3	2-AA	9-AA	2-AA	NQO	2-AA	NaN3	2-AA
Total Plates	190	190	90	90	88	88	152	152	190	200
Number of Values	89	89	39	39	38	38	70	70	89	94
Minimum	89	201	546	138	120	106	222	126	195	450
Maximum	1197	1544	1562	352	2313	588	2613	737	2289	2229
Mean	362	769	867	262	977	378	1606	394	1360	1365
Standard Deviation	201.9	257.4	172.3	51.8	429.9	142.3	488.8	142.2	305.0	314.9

Conclusions:

The test substance was considered to be non-mutagenic with and without metabolic activation in bacteria.

Executive summary:

The test item was examined for its mutagenic activity in two series of a bacterial reverse mutation assay (OECD 471) employing Salmonella typhimurium TA 98, TA 100, TA 1535 and TA 1537, and Escherichia coli WP2 uvrA as indicator organisms.

The plate incorporation test with and without addition of liver S9 mix from Aroclor 1254-pre-treated rats was used. In this study, two experimental series were performed. In the experiments with S9 mix, 10 % and 30% S9 were used in the first and second series, respectively.

Treatments of all tester strains were performed using the test item solved in ultrapure water in the absence and in the presence of S9 mix, using final concentrations between 5 and 5000 µg/plate, plus vehicle and positive controls.

The strain specific positive control test materials, namely daunomycin, sodium azide, 4-nitroquin-olin-N-oxide, and 9-aminoacridine in the absence of S9 mix, yielded the expected mutant frequencies that were greatly in excess of the negative controls. The genotype of the tester strains used was thus confirmed. 2-Aminoanthracene which requires metabolic activation, was strongly mutagenic. This indicates that the exogenous metabolizing system used in the present investigation (S9 mix) was functioning. Thus, the study was considered valid.

No test material precipitate was observed on the plates at any of the doses tested in either the presence or absence of S9-mix. During the experiments there was no evidence of toxicity to the bacteria caused by the test material exposure.

Under the conditions described, there were no relevant increases in revertant numbers after treatment with the test item observed in both, the absence and presence of S9 mix.

In conclusion, the test material was considered to be non-mutagenic under the described experimental conditions.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Genetic toxicity in vivo

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Bacterial Reverse Mutation Test

The test item was examined for its mutagenic activity in two series of a bacterial reverse mutation assay (OECD 471) employing Salmonella typhimurium TA 98, TA 100, TA 1535 and TA 1537, and Escherichia coli WP2 uvrA as indicator organisms.

The plate incorporation test with and without addition of liver S9 mix from Aroclor 1254-pre-treated rats was used. In this study, two experimental series were performed. In the experiments with S9 mix, 10 % and 30% S9 were used in the first and second series, respectively.

Treatments of all tester strains were performed using the test item solved in ultrapure water in the absence and in the presence of S9 mix, using final concentrations between 5 and 5000 µg/plate, plus vehicle and positive controls.

The strain specific positive control test materials, namely daunomycin, sodium azide, 4-nitroquin-olin-N-oxide, and 9-aminoacridine in the absence of S9 mix, yielded the expected mutant frequencies that were greatly in excess of the negative controls. The genotype of the tester strains used was thus confirmed. 2-Aminoanthracene which requires metabolic activation, was strongly mutagenic. This indicates that the exogenous metabolizing system used in the present investigation (S9 mix) was functioning. Thus, the study was considered valid.

No test material precipitate was observed on the plates at any of the doses tested in either the presence or absence of S9-mix. During the experiments there was no evidence of toxicity to the bacteria caused by the test material exposure.

Under the conditions described, there were no relevant increases in revertant numbers after treatment with the test item observed in both, the absence and presence of S9 mix.

In conclusion, the test material was considered to be non-mutagenic under the described experimental conditions.

Justification for classification or non-classification

Classification, Labeling, and Packaging Regulation (EC) No 1272/2008

The available experimental test data are reliable and suitable for classification purposes under Regulation (EC) No 1272/2008. The results indicate that the substance is non-mutagenic. Based on available data on genetic toxicity, the test item is not classified according to Regulation (EC) No 1272/2008 (CLP), as amended for the tenth time in Regulation (EC) No 2017/776.