Zinc dodecyl hydrogen disulphate

Endpoint summary

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

Bacterial reverse mutation assay (Ames / OECD guideline 471): negative
In vitro mammalian cell gene mutation assay (MLA / OECD guideline 476): negative

Link to relevant study records

Reference

Reference 1

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Justification for type of information:

Refer to the Analogue Approach Justification document provided in IUCLID Section 13.

Reason / purpose for cross-reference:

read-across source

Statistics:

No.

Key result

Species / strain:

S. typhimurium TA 1535

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

at 1500 µg/plate

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

Key result

Species / strain:

S. typhimurium TA 1537

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

at 1500 µg/plate

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

Key result

Species / strain:

S. typhimurium TA 98

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

at 1500 µg/plate

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

Key result

Species / strain:

S. typhimurium TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

at 1500 µg/plate

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

Key result

Species / strain:

S. typhimurium TA 1538

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

at 1500 µg/plate

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

Conclusions:

C12-14AS Zn is not considered to be mutagenic in bacteria with or with metabolic activation.

Reference 2

Endpoint:

in vitro gene mutation study in mammalian cells

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Justification for type of information:

Refer to the Analogue Approach Justification document provided in IUCLID Section 13.

Reason / purpose for cross-reference:

read-across source

Key result

Species / strain:

mouse lymphoma L5178Y cells

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

-S9: 70, 80 and 90 µg/mL; +S9: 95 µg/mL

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

Positive controls validity:

valid

RESULTS OF EXPERIMENTS 1-8

Eight acceptable experiments were conducted, five in the absence of S9 mix.

In the first of these, statistically significant increases in mutant fraction were observed at three dose levels: 6.25, 25, and 50 µg/mL; 100 µg/mL was a lethal concentration in cells (see Table 1). Over the nonlethal range, there were generally elevated mutant fractors, the highest being 1.9-fold the control level at 25 µg/mL. Although these increases in mutant fraction were significant, the lack of an obvious dose-related response with a relatively soluble chemical over a dose range which was not toxic encouraged speculation that the increases were not due to treatment with the test material.

 

Table 1. Experiment 1 - 4 h exposure - Without Metabolic Activation

Concentration [µg/mL]	Cloning efficiency	Relative Total Growth	Mutants per 1E+06 surviving cells	Mutation factor	Average Mutation factor
DMSO (NC)	62	95	56	30	43
	68	98	90	44
	65	95	66	34
	80	112	153	64
3.125	77	106	84	36	61
	59	97	151	85
6.25	71	119	176	83	78*
	73	107	160	74
12.5	90	145	133	49	65
	67	133	160	80
25	84	90	247	99	83*
	65	88	130	67
50	86	82	192	75	69*
	76	98	145	64
100	lethal	lethal	n.a.	n.a.	n.a.
			n.a.	n.a.
MMS (15 µg/mL) PC	27	21	135	167	232*
	25	23	219	298

MMS = methylmethanesulfonate; NC = negative control; PC = positive control; *p < 0.05; n.a. = not applicable

 

In the second experiment without S9 mix, there was a clearly significant response at 60 µg/mL, but at no other concentration. The RTG was about 22%

Table 2. Experiment II - 4 h exposure - Without Metabolic Activation

Concentration [µg/mL]	Cloning efficiency	Relative Total Growth	Mutants per 1E+06 surviving cells	Mutation factor	Average Mutation factor
DMSO (NC)	80	100	115	48	48
	74	108	119	53
	86	106	115	45
	61	87	83	45
10	75	112	107	47	58
	66	94	137	69
20	67	89	119	59	52
	53	76	70	44
30	71	70	98	46	60
	64	86	144	75
40	77	77	126	54	n.a.
50	94	61	191	68	74
	68	56	163	80
60	81	27	365	150	203*
	77	16	595	256
70	lethal	lethal	n.a.	n.a.	n.a.
			n.a.	n.a.
MMS (15 µg/mL) PC	34	30	683	666	664*
	29	27	573	662

MMS = methylmethanesulfonate; NC = negative control; PC = positive control; *p < 0.05; n.a. = not applicable

Experiment 3 gave a statistically significant response (1.7-fold increase) at 60 µg/mL, but not at the next higher concentration of 65 µg/mL. The mutant fraction at 70 µg/ml was only 44/106 survivors, so this single culture result supported the view that the statistically significant result at the lower dose level was a chance event. Thus, this experiment was judged to be questionable.

 

Table 3. Experiment 3 - 4 h exposure - Without Metabolic Activation

Concentration [µg/mL]	Cloning efficiency	Relative Total Growth	Mutants per 1E+06 surviving cells	Mutation factor	Average Mutation factor
DMSO (NC)	76	103	84	37	34
	71	104	73	34
	82	97	98	40
	74	96	58	26
50	60	71	65	36	36
	75	75	82	36
55	68	44	68	33	38
	53	71	69	44
60	72	63	107	50	56*
	72	74	134	62
65	64	71	87	45	42
	79	68	93	39
70	80	83	107	44	n.a.
MMS (15 µg/mL) PC	36	26	127	119	158*
	26	23	156	197

MMS = methylmethanesulfonate; NC = negative control; PC = positive control; *p < 0.05; n.a. = not applicable

 

However, the succeeding experiments 4 and 5 without S9 mix were unambiguously negative; therefore the test substance was considered to be non-mutagenic in the absence of S9 mix.

Two experiments (6 and 7) were performed in the presence of S9 mix, showing unambiguously negative results. The last experiment with S9 mix was inconclusive because the cloning efficiency at 80 µg/mL was about 86% and there was no indication of a mutagenic response. However, based on the two experiments with S9 mix showing clearly negative results, the test substance was considered to be not mutagenic in the presence of S9 mix.

Conclusions:

C12-14AS Zn was not considered to be mutagenic, neither in the presence nor in the absence of metabolic activation.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Genetic toxicity in vivo

Description of key information

Mammalian Bone Marrow Chromosome Aberration Test (CA / OECD guideline 475): negative

Mammalian Erythrocyte Micronucleus Test (MNT / OECD guideline 474): negative

Link to relevant study records

Reference

Reference 1

Endpoint:

in vivo mammalian somatic cell study: cytogenicity / bone marrow chromosome aberration

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

weight of evidence

Justification for type of information:

Refer to the Analogue Approach Justification document provided in Section 13

Reason / purpose for cross-reference:

read-across source

Key result

Sex:

male/female

Genotoxicity:

negative

Toxicity:

no effects

Vehicle controls validity:

valid

Negative controls validity:

not applicable

Positive controls validity:

valid

Conclusions:

C12-14AS Zn was not considered to be produce chromosome aberrations in vivo.

Reference 2

Endpoint:

in vivo mammalian somatic cell study: cytogenicity / erythrocyte micronucleus

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

weight of evidence

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

According to the ECHA guidance document “Practical guide 6: How to report read-across and categories (March 2010)”, the reliability was changed from RL1 to RL2 to reflect the fact that this study was conducted on a read-across substance.

Justification for type of information:

Refer to the Analogue Approach Justification document provided in Section 13

Reason / purpose for cross-reference:

read-across source

Key result

Sex:

male/female

Genotoxicity:

negative

Toxicity:

no effects

Vehicle controls validity:

valid

Negative controls validity:

not applicable

Positive controls validity:

valid

Conclusions:

Interpretation of results: negative
C12-14AS Zn was not considered to be clastogenic.

Executive summary:

No increase in micronucleated polychromatic erythrocytes was observed in this micronucleus test. Therefore, the test substance was not considered to be clastogenic.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Additional information

The lack of mutagenic activity for the alkyl sulfate analogues is predictable based on structural and mechanistic considerations. Mutagens are chemicals that either 1) contain highly reactive electrophilic centers capable of interacting with nucleophilic sites on DNA (direct acting agents) or 2) can be metabolized to highly reactive electrophiles. The chemical structures represented by this chemical class do not contain electrophilic functional groups or functional groups capable of being metabolized to electrophiles. Alkyl sulfates with fully saturated carbon chains are not metabolized to reactive electrophiles. The consistent lack of mutagenic activity with alkyl sulfates is consistent with these mechanistic predictions.

There is no study regarding genotoxicity available for C12-14 AS Zn (CAS n.a.). Therefore this endpoint is covered by read across to structurally related alkyl sulfates (AS) for weight-of-evidence approach, i.e. C12-14 AS Na (CAS 85586-07-8), C12 AS Na (CAS 151-21-3), C12-15 AS Na (CAS 68890-70-0) and C12-14 AS TEA (CAS 90583-18-9). The possibility of a read-across to other alkyl sulfates in accordance with Regulation (EC) No 1907/2006 Annex XI 1.5 “Grouping of substances and read-across approach” was assessed. In Annex XI 1.5 it is given that a read-across approach is possible for substances whose physicochemical, toxicological and ecotoxicological properties are likely to be similar or follow a regular pattern as a result of structural similarity. The AS reported within the AS analogues show structural similarity. The most important common structural feature of the members of the analogue approach is the presence of a predominantly linear aliphatic hydrocarbon chain with a polar sulfate group, neutralized with a counter ion. This structural feature confers the surfactant properties of the alkyl sulfates. The surfactant property of the members of the AS analogues in turn represent the predominant attribute in mediating effects on mammalian health. Therefore, the AS of the AS analogues have similar physicochemical, environmental and toxicological properties, validating the read-across approach. The approach of grouping different AS for the evaluation of their effects on human health and the environment was also made by the OECD in the SIDS initial assessment profile [1] and by a voluntary industry program carrying out Human and Environmental Risk Assessments (HERA [2]), further supporting the read across approach between structurally related AS.

Zinc compounds are recognised as zinc category in the OECD HPV program [3]. The zinc category includes six compounds (zinc metal, zinc oxide, zinc distearate, zinc chloride, zinc sulphate, and trizinc bis(orthophosphate). Available data show that zinc salts have moderate effects upon acute oral application and may be irritating to skin and eyes. No genotoxic potential in vivo is reported for the above mentioned zinc salts. Therefore, contribution of zinc to the endpoint genotoxicity is considered to be negligible when assessing human health effects of C12-14AS Zn (CAS n.a.). 

Mutagenicity in bacteria

Mutagenicity in bacteria was assessed in a study performed according to OECD Guideline 471. Tester strain TA 102 or E.coli were not used during the conduct of the study (Kao, 1996). In this study with C12-14 AS Na (CAS 85586-07-8), Salmonella typhimurium strains TA 1535, TA 1537, TA 98, TA 1538 and TA 100 were treated with and without addition of a rat liver S9-mix. The dose range was 5, 15, 50, 150, 500, and 1500 µg/plate without and 5, 15, 50, 150, 500, 1500 and 5000 µg/plate with metabolic activation in the first experiment as well as 5, 15, 50, 150, 500, and 1500 µg/plate (with and without S9 mix) in the second experiment. Results achieved with vehicle (DMSO) and positive controls were valid. Cytotoxicity was observed in presence and absence of metabolic activation at 1500 µg/plate while no genotoxicity was observed.

 

Mutagenicity in mammalian cells

The mutagenicity of C12 AS Na (CAS 151-21-3) in a mammalian cell line was investigated similar to OECD guideline 476 using the mouse lymphoma L5178Y cells with and without metabolic activation (McGregor, 1988). The test concentrations were 3.125, 6.25, 10, 12.5, 20, 25, 30, 40, 50, 55, 60, 65, 70, 80 and 100 µg/mL without and 50, 55, 60, 65, 70, 75, 80, 85, 90 and 95 µg/mL with metabolic activation. Results achieved with the negative (untreated), vehicle (DMSO) and positive controls were valid. Cytotoxicity was observed in presence and absence of metabolic activation while no genotoxicity was observed under both circumstances.

 

Clastogenicity in vivo

The potential of C12-14 AS TEA (CAS 90583-18-9, analytical purity approx. 41%) to induce micronuclei in vivo was assessed in a study conducted according to OECD guideline 474 with CFW-1 mouse (BASF, 1987). The test substance was administered via gavage at doses of 400, 2000 and 4000 mg/kg bw to 7 animals per sex and dose. Bone marrow was sampled 24 h (400 and 2000 mg/kg bw) and 24, 48 and 72 h (4000 mg/kg bw) after gavage. Results achieved with the vehicle (DMSO) and positive controls were valid. No signs of toxicity were noted. As no enhanced chromosome aberrations were observed in this micronucleus test the test substance was considered to be not clastogenic.

The potential of C12-15 AS Na (CAS 68890-70-0, analytical purity approx. 30%) to induce in vivo chromosomal aberration was assessed in a study conducted similar to OECD guideline 475 in rats (Unilever, 1976). The test substance was administered via feed at a dose of 1.13% for a period of 90 days to 6 animals per sex and dose and bone marrow was sampled thereafter. Results achieved with the vehicle (DMSO) and positive controls were valid. No signs of toxicity were noted. As no enhanced chromosome aberrations were observed in this chromosomal aberration test the test substance was considered to be not clastogenic.

In conclusion, the read across substances did not show any genotoxic potential. This is supported by the conclusions of the HERA Draft report “AS are not genotoxic, mutagenic or carcinogenic…” and the conclusions of the SIDS initial assessment profile “Alkyl sulfates of different chain length and with different counter ions were not mutagenic in standard bacterial and mammalian cell systems [...]. There was also no indication for a genotoxic potential of alkyl sulfates in various in vivo studies on mice […].”

 

[1] SIDS initial assessment profile, (2007); http://www.aciscience.org/docs/Alkyl_Sulfates_Final_SIAP.pdf

[2] (HERA Draft report, 2002); http://www.heraproject.com/files/3-HH-04-%20HERA%20AS%20HH%20web%20wd.pdf

[3] SIDS initial assessment profile, (2005);http://webnet.oecd.org/Hpv/UI/handler.axd?id=fddec5fa-9727-413a-9d67-41c2154cd362

Justification for classification or non-classification

The available data on genetic toxicity do not meet the criteria for classification according to Regulation (EC) 1272/2008 or Directive 67/548/EEC, and are therefore conclusive but not sufficient for classification.