Hexadecylamine

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Data source

Reference

Reference Type:

publication

Title:

Unnamed

Year:

1988

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Principles of method if other than guideline:

Data from summary document with only 4 strains; acceptable because data consistent with other data in subcategory. The endpoint has been adequately characterized. (American Chemistry Council, Fatty Nitrogen Derivatives Panel, Amines Task Group).

GLP compliance:

not specified

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

1

Reference substance name:

Hexadecylamine

EC Number:

205-596-8

EC Name:

Hexadecylamine

Cas Number:

143-27-1

Molecular formula:

C16H35N

IUPAC Name:

hexadecan-1-amine

Test material form:

solid

Method

Target gene:

Histidine (S. typhimurium)

Species / strain

Species / strain 1

Species / strain / cell type:

S. typhimurium TA 97

Species / strain 2

Species / strain / cell type:

S. typhimurium TA 98

Species / strain 3

Species / strain / cell type:

S. typhimurium TA 100

Species / strain 4

Species / strain / cell type:

S. typhimurium TA 1535

Metabolic activation:

with and without

Metabolic activation system:

The S-9 (9,000g supernatant) fractions of Aroclor 1254-induced, male Sprague-Dawley rat and male Syrian hamster livers were prepared as described previously [Haworth et al, 1983]. The S-9 mixes were prepared immediately prior to use and contained either 10% or 30% S-9. The test substance was tested in the absence of metabolic activation and with rat and hamster S-9 fractions.

Test concentrations with justification for top dose:

0.3, 1.0, 3, 10, 33, 100 µg/plate based on toxicity pretest

Vehicle / solvent:

DSMO

Controls

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

9-aminoacridine

sodium azide

other: 4-nitro-o-phenylenediamine: TA98 (without S9); 2-aminoan-thracene: all strains (with S9)

Details on test system and experimental conditions:

The preincubation assay was performed as described previously [Haworth et al., 1983], with some differences, as described below. The test chemical (0.05 ml), Salmonella culture (0.10 ml), and S-9 mix or buffer (0.50 ml) were incubated at 37°C, without shaking, for 20 min. Chemicals known or suspected to be volatile were incubated in capped tubes. The top agar was added and the contents of the tubes were mixed and poured onto the surface of petri dishes containing Vogel-Bonner medium [Vogel and Bonner, 1956]. The histidine-independent (his+) colonies arising on these plates were counted following two days incubation at 37°C. Plates were machine counted (New Brunswick, Edison, NJ; Artek, Farmingdale, NY) unless precipitate was present which interfered with the count, or the color of the test chemical on the plate reduced the contrast between the colonies and the background agar. At the discretion of the investigators, plates with low numbers of colonies were counted by hand.
Variations in the protocol among the tested chemicals reflect the evolution of the protocol originally described by Haworth et al. [1983].
The test substance was tested initially in a toxicity assay to determine the appropriate dose range for the mutagenicity assay. The toxicity assay was performed using TA100 or the sytem developed by Waleh et al. [1982]. Toxic concentrations were those that produced a decrease in the number of his+ colonies, or a clearing in the density of the background lawn, or both. The test substance was tested initially at half-log dose intervals up to a dose that elicited toxicity, or to a dose immediately below one which was toxic in the preliminary toxicity test. Subsequent trials occasionally used narrower dose increments and may not have included doses in the toxic range. Concurrent solvent and positive controls were run. The positive controls in the absence of metabolic activation were sodium azide (TA1535 and TA100), 9-aminoacridine (TA97 and TA1537), and 4-nitro-o-phenylenediamine (TA98). The positive control for metabolic activation with all strains was 2-aminoan-thracene.

Evaluation criteria:

The data were evaluated as described previously [Zeiger et al., 1987]. Evaluations were made at both the individual trial and overall chemical levels. Individual trials were judged mutagenic (+), weakly mutagenic (+W), questionable (?), or nonmutagenic (-), depending on the magnitude of the increase of his+ revertants, and the shape of the dose-response. A trial was considered questionable (?) if the dose-response was judged insufficiently high to support a call of " +W," if only a single dose was elevated over the control, or if the increase seen was not dose related. The distinctions between a questionable mutagenic response and a nonmutagenic or weak mutagenic response and between a weak mutagenic response and mutagenic response are highly subjective. It was not necessary for a response to reach twofold over background for a chemical to be judged mutagenic. A chemical was judged mutagenic (+) or weakly mutagenic (+W) if it produced a reproducible dose-related response over the solvent control in replicate trials. A chemical was judged questionable (?) if the results of individual trials were not reproducible, if increases in his4" revertants did not meet the criteria for a "+W" response, or if only single doses produced increases in his +revertants in repeat trials. Chemicals were judged nonmutagenic (-) if they did not meet the criteria for a mutagenic or questionable response. The chemicals were decoded by the chemical repository only after a determination had been made regarding their mutagenicity or nonmutagenicity.

Results and discussion

Test results

Key result

Species / strain:

S. typhimurium, other: TA 97, 98, 100, 1535

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

at 100 µg/plate

Vehicle controls validity:

valid

True negative controls validity:

not examined

Positive controls validity:

valid

Additional information on results:

No historical control data is available, but the control plates without activation were comparable to that described in literature.

Any other information on results incl. tables

Table 1: Detailed results for TA100 and TA1535

Dose	TA100										TA1535
	NA		10% HLI		30% HLI		10% RLI		30% RLI		NA		10% HLI		30% HLI		10% RLI		30% RLI
µg/PLATE	mean	SD	mean	SD	mean	SD	mean	SD	mean	SD	mean	SD	mean	SD	mean	SD	mean	SD	mean	SD
0.000	120	3.1	108	1.3	127	7.9	118	4.4	142	1.9	10	3.4	6	2.2	16	1.9	6	0.9	9	2.6
0.300	106	3.1									15	4.3
1.000	93	5.5	113	28.5	131	2.9	130	33.5	127	17.1	17	2.5	12	5.8	13	0.7	8	2.8	17	4.0
3.000	63	4.2	110	17.3	117	10.5	119	8.5	110	9.9	11	3.2	6	0.7	7	1.3	7	1.5	7	1.7
10.000	73	6.8	121	6.9	131	5.2	165	15.0	113	12.7	15	0.9	7	1.0	9	3.0	7	1.0	12	1.9
33.000	69	9.8	116	15.0	124	9.0	161	12.5	114	9.3	12	2.3	10	1.5	9	0.3	8	3.2	8	1.9
100.000			78	10.0	112	7.2	97	5.5	134	13.7			8	3.0	5	1.3	4	0.9	9	0.9
POS	274	5.2	2127	115.0	748	9.6	1380	84.1	352	4.6	186	3.0	635	8.0	534	33.8	460	24.6	110	11.1

Abbreviations:

POS: positive control

NA: not activated

HLI: Aroclor 1254-induced hamster liver S-9

RLI: Aroclor 1254-induced rat liver S-9

 

Table 2: Detailed results of TA97 and TA98

Dose	TA97										TA98
	NA		10% HLI		30% HLI		10% RLI		30% RLI		NA		10% HLI		30% HLI		10% RLI		30% RLI
µg/PLATE	mean	SD	mean	SD	mean	SD	mean	SD	mean	SD	mean	SD	mean	SD	mean	SD	mean	SD	mean	SD
0.000	153	2.6	216	5.2	173	9.2	216	15.6	224	7.6	12	3.7	45	5.7	22	2.5	34	1.7	21	0.9
0.300	155	2.6									11	1.3
1.000	140	9.8	255	19.7	199	30.0	255	11.8	234	12.2	14	3.2	39	6.9	22	2.3	35	3.5	28	0.9
3.000	126	9.0	230	3.3	183	15.9	235	19.3	191	13.3	7	0.6	35	11.2	21	1.5	28	1.5	25	2.8
10.000	114	6.1	255	11.3	191	16.3	286	11.3	207	14.3	10	1.5	36	2.0	15	2.0	31	4.7	15	2.5
33.000	101	4.0	225	9.0	182	12.3	265	4.2	204	11.3	9	2.3	32	2.0	13	3.4	26	0.9	19	4.0
100.000			195	14.2	192	5.2	212	15.0	203	9.6			21	4.1	13	3.5	22	6.0	23	3.5
POS	106	141.6	1121	53.1	1007	50.1	1767	80.4	579	7.0	740	9.8	1207	133.9	234	7.0	830	110.3	151	3.7

Abbreviations:

POS: positive control

NA: not activated

HLI: Aroclor 1254-induced hamster liver S-9

RLI: Aroclor 1254-induced rat liver S-9

Applicant's summary and conclusion

Conclusions:

Based on the test results it can be stated that the test item is not mutagenic in the strains TA 100, TA 1535, TA 97 and TA 98 of Salmonella typhimurium with and without metabolic activation.

Executive summary:

The test item was tested for mutagenicity with the strains TA 100, TA 1535, TA97 and TA 98 of Salmonella typhimurium. The mutagenicity study was conducted in the absence and in the presence of a metabolizing system derived from rat and hamster liver homogenate. Control plates without mutagen showed that the number of spontaneous revertant colonies was comparable to that described in literatur. All the positive control compounds gave the expected increase in the number of revertant colonies. In the cytotoxicity trials, the test compound proved to be toxic to most of the bacterial strains at 100 microgram/plate. These tests demonstrated that in the presence or absence of the metabolic activation system the test compound did not show a dose dependent increase in the number of revertants in any of the bacterial strains. Summarizing, it can be stated that the test item is not mutagenic in these bacterial test systems either with or without exogenous metabolic activation at the dose levels investigated.