Aluminium, 4,5-dihydro-5-oxo-1-(4-sulfophenyl)-4-[(4-sulfophenyl)azo]-1H-pyrazole-3-carboxylic acid complex

Endpoint summary

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

Pigment yellow 100 was studied for its ability to induce mutations in strains of Salmonella typhimurium.

 

The test compound was dissolved in DMSO and was tested at concentration of 0, 100, 333, 1000, 3333, 6666 or 10000 µg/plate using Salmonella typhimurium TA100, TA1535, TA1537, TA97 and TA98 in the presence and absence of 10 % and 30 % rat and hamster liver S9 metabolic activation system. Preincubation assay was performed with a preicubation for 20 mins. The plates were observed for histidine independence after 2 days incubation period. Concurrent solvent and positive controls were included in the study.

 

Pigment yellow 100 is not mutagenic to the Salmonella typhimurium TA100, TA1535, TA1537, TA97 and TA98 in the presence and absence of rat and hamster liver S9 metabolic activation system  and hence it is not likely to classify as a gene mutant in vitro.

Link to relevant study records

Reference

Endpoint:

in vitro gene mutation study in bacteria

Remarks:

Type of genotoxicity: gene mutation

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

data from handbook or collection of data

Justification for type of information:

Data is from peer reviewed publication

Qualifier:

according to guideline

Guideline:

other: Refer below principle

Principles of method if other than guideline:

The test chemical Pigment yellow 100 (C.I. 19140) was studied for its ability to induce mutations in strains of Salmonella typhimurium.

GLP compliance:

not specified

Type of assay:

bacterial reverse mutation assay

Specific details on test material used for the study:

- Name of the test material: Pigment yellow 100
- IUPAC name: aluminum tris(4-{[3-carboxy-5-oxo-1-(4-sulfophenyl)-4,5-dihydro-1H-pyrazol-4-yl]diazenyl}benzenesulfonate)
- Molecular formula: C48H33AlN12O27S6
- Molecular weight: 495.4038 g/mol
- Substance type: Organic
- Purity: Labelled: 26%; Analyzed: 27%
- Inchi: 1S/C16H12N4O9S2.Al/c21-15-13(18-17-9-1-5-11(6-2-9)30(24,25)26)14(16(22)23)19-20(15)10-3-7-12(8-4-10)31(27,28)29;/h1-8,13H,(H,22,23)(H,24,25,26)(H,27,28,29);/b18-17+;

Target gene:

Histidine

Species / strain / cell type:

S. typhimurium, other: TA100, TA1535, TA1537, TA97 and TA98

Details on mammalian cell type (if applicable):

Not applicable

Additional strain / cell type characteristics:

not specified

Cytokinesis block (if used):

No data

Metabolic activation:

with and without

Metabolic activation system:

10% and 30% HLI and RLI S-9 (9,000 g supernatant) fractions were prepared from Aroclor 1254-induced, male Sprague- Dawley rat and male Syrian hamster livers

Test concentrations with justification for top dose:

0, 100, 333, 1000, 3333, 6666 or 10000 µg/plate

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: Dimethyl Sulfoxide (DMSO)
- Justification for choice of solvent/vehicle: The test chemical was soluble in DMSO

Untreated negative controls:

not specified

Negative solvent / vehicle controls:

yes

Remarks:

DMSO

True negative controls:

not specified

Positive controls:

yes

Positive control substance:

other: 2-aminoanthracene

Remarks:

For strains tested with S9

Untreated negative controls:

not specified

Negative solvent / vehicle controls:

yes

Remarks:

DMSO

True negative controls:

not specified

Positive control substance:

sodium azide

Remarks:

For strains TA100 and TA1535 tested in the absence of S9

Untreated negative controls:

not specified

Negative solvent / vehicle controls:

yes

Remarks:

DMSO

True negative controls:

not specified

Positive controls:

yes

Positive control substance:

9-aminoacridine

Remarks:

For strains TA97 and TA1537 tested in the absence of S9

Untreated negative controls:

not specified

Negative solvent / vehicle controls:

yes

Remarks:

DMSO

True negative controls:

not specified

Positive controls:

yes

Positive control substance:

other: 4-nitro-o-phenylenediamine

Remarks:

For strain TA98 tested in the absence of S9

Details on test system and experimental conditions:

METHOD OF APPLICATION: preincubation

DURATION
- Preincubation period: 20 mins
- Exposure duration: 48 hrs
- Expression time (cells in growth medium): 48 hrs
- Selection time (if incubation with a selection agent): No data available
- Fixation time (start of exposure up to fixation or harvest of cells): No data available

SELECTION AGENT (mutation assays): No data available
SPINDLE INHIBITOR (cytogenetic assays): No data available
STAIN (for cytogenetic assays): No data available

NUMBER OF REPLICATIONS: Triplicate

NUMBER OF CELLS EVALUATED: No data available

DETERMINATION OF CYTOTOXICITY
- Method: mitotic index; cloning efficiency; relative total growth; other: No data available

OTHER EXAMINATIONS:
- Determination of polyploidy: No data available
- Determination of endoreplication: No data available
- Other: No data available

OTHER: No data available

Rationale for test conditions:

No data

Evaluation criteria:

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 two fold 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 reponse over the solvent control in replicate trials. A chemical was judged questionable (?) if the results of individual trials were not reproducible, if increases in his+ 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.

Statistics:

Mean ± SD

Species / strain:

S. typhimurium, other: TA100, TA1535, TA1537, TA97 and TA98

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

valid

Untreated negative controls validity:

not specified

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
- Effects of pH: No data available
- Effects of osmolality: No data available
- Evaporation from medium: No data available
- Water solubility: No data available
- Precipitation: No data available
- Other confounding effects: No data available

RANGE-FINDING/SCREENING STUDIES: All chemicals were tested initially in a toxicity assay to determine the appropriate dose range for the mutagenicity assay. The toxicity assay was performed using TA100. 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.

COMPARISON WITH HISTORICAL CONTROL DATA: Yes

ADDITIONAL INFORMATION ON CYTOTOXICITY: No data available

Remarks on result:

other: No mutagenic potential

Strain: TA100

Dose	No Activation  (Negative)		No Activation  (Negative)		30% RLI  (Negative)		30% HLI  (Negative)		10% RLI  (Negative)		10% HLI  (Negative)
Protocol	Preincubation		Preincubation		Preincubation		Preincubation		Preincubation		Preincubation
ug/Plate	Mean	±SEM	Mean	±SEM	Mean	±SEM	Mean	±SEM	Mean	±SEM	Mean	±SEM
0	146	10	127	7.5	145	14.7	167	6.8	107	10.1	107	6.8
100	133	3.7	137	7.5	143	4.7	165	17.8	123	16.5	117	11.4
333	130	3.8	152	5	116	5	155	10.8	122	4.4	128	8.7
1000	146	6.1	136	6.7	127	.7	116	16.5	130	13.5	117	13.1
3333	109	4	65	7.9	130	15.3	118	5.9	106	5.8	85	8.7
6666			16	.7					75	11.4	53	8.9
10000	37	9			65	8.2	61	12.6
Positive Control	604	4	550	21.1	478	14.7	529	14.2	515	46	548	52.7

Strain: TA1535

Dose	No Activation  (Negative)		No Activation  (Negative)		30% RLI  (Negative)		30% HLI  (Negative)		10% RLI  (Negative)		10% HLI  (Negative)
Protocol	Preincubation		Preincubation		Preincubation		Preincubation		Preincubation		Preincubation
ug/Plate	Mean	±SEM	Mean	±SEM	Mean	±SEM	Mean	±SEM	Mean	±SEM	Mean	±SEM
0	31	1	20	.6	15	1.8	9	2	8	.6	10	1.5
100	26	1.5	20	1.5	14	1.9	10	1.8	11	1.2	12	2
333	30	5.2	16	.6	16	2.2	8	1.2	7	2.5	7	.9
1000	30	4	25	6.9	15	1.9	8	1.5	9	.9	8	.9
3333	24	3.7	15	2.5	11	1.2	8	3.1	9	2.7	6	1.5
6666			14	2.6					6	.9	6	.3
10000	21	1.9			13	1.9	7	.7
Positive Control	357	22.3	499	21.5	218	6.6	388	17.1	121	8.4	219	4.6

Strain: TA1537

Dose	No Activation  (Negative)		30% RLI  (Negative)		30% HLI  (Negative)
Protocol	Preincubation		Preincubation		Preincubation
ug/Plate	Mean	±SEM	Mean	±SEM	Mean	±SEM
0	8	.7	14	1.5	15	.6
100	6	1.5	11	1.7	12	3.5
333	10	1.9	15	1.8	12	1.9
1000	11	1.8	12	.6	10	.3
3333	6	.6	9	1.5	6	0
10000	5	1.2	8	1.9	4	0
Positive Control	309	5	41	2.3	46	3.3

Conclusions:

Pigment yellow 100 is not mutagenic to the Salmonella typhimurium TA100, TA1535, TA1537, TA97 and TA98 in the presence and absence of rat and hamster liver S9 metabolic activation system and hence it is not likely to classify as a gene mutant in vitro.

Executive summary:

Pigment yellow 100 was studied for its ability to induce mutations in strains of Salmonella typhimurium.

 

The test compound was dissolved in DMSO and was tested at concentration of 0, 100, 333, 1000, 3333, 6666 or 10000 µg/plate using Salmonella typhimurium TA100, TA1535, TA1537, TA97 and TA98 in the presence and absence of 10 % and 30 % rat and hamster liver S9 metabolic activation system. Preincubation assay was performed with a preicubation for 20 mins. The plates were observed for histidine independence after 2 days incubation period. Concurrent solvent and positive controls were included in the study.

 

Pigment yellow 100 is not mutagenic to the Salmonella typhimurium TA100, TA1535, TA1537, TA97 and TA98 in the presence and absence of rat and hamster liver S9 metabolic activation system  and hence it is not likely to classify as a gene mutant in vitro.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Genetic toxicity in vivo

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Gene mutation in vitro:

Data available for the target chemical and its reas across have been reviewed to determine the mutagenic nature of aluminum tris(4-{[3-carboxy-5-oxo-1-(4-sulfophenyl)-4,5-dihydro-1H-pyrazol-4-yl]diazenyl}benzenesulfonate). The studies are as mentioned below:

Pigment yellow 100 (CAS no 12225 -21 -7) was studied by Zeiger et al (Environmental and Molecular Mutagenesis, 1998) for its ability to induce mutations in strains of Salmonella typhimurium. The test compound was dissolved in DMSO and was tested at concentration of 0, 100, 333, 1000, 3333, 6666 or 10000 µg/plate using Salmonella typhimurium TA100, TA1535, TA1537, TA97 and TA98 in the presence and absence of 10 % and 30 % rat and hamster liver S9 metabolic activation system. Preincubation assay was performed with a preicubation for 20 mins. The plates were observed for histidine independence after 2 days incubation period. Concurrent solvent and positive controls were included in the study. Pigment yellow 100 is not mutagenic to the Salmonella typhimurium TA100, TA1535, TA1537, TA97 and TA98 in the presence and absence of rat and hamster liver S9 metabolic activation system  and hence it is not likely to classify as a gene mutant in vitro.

The above data for the target chemical is further supported by the data from read across chemical.

In a study performed by Das and Mukherjee (Human Genetics, 2004) Ames mutagenicity assay was performed to evaluate the mutagenic nature of the structurally and functionally similar read across chemical tartrazine (RA CAS no 1934 -21 -0; IUPAC name: Trisodium 5-hydroxy-1-(4-sulphophenyl)-4-(4-sulphophenylazo)pyrazole-3-carboxylate) using the plate incorporation assay. Tartrazine was dissolved in sterile double distilled water and was tested at a concentration of 0, 100, 250, 500 and 1000 μg /plate. The plates were inverted within an hour and placed in a dark vented incubator at 37⁰C for 48 hours. Positive controls (for TA97a and TA98, 20 μg/plate nitro phenylene diamine and for TA100, 1.5 μg/plate sodium azide) and negative controls were maintained concurrently for all the experiments. Three plates were used for each set. After 48 hours of incubation, the revertant colonies were counted. Tartrazine did not induce gene mutation in Salmonella typhimurium strains TA97a, TA98 and TA100 and hence it is not likely to classify as a gene mutant in vitro.

Chromosomal aberration test was performed by Ishidate et al (Food and chemical toxicology, 1988) for the structurally and functionally similar read across chemical tartrazine (RA CAS no 1934 -21 -0; IUPAC name:Trisodium 5-hydroxy-1-(4-sulphophenyl)-4-(4-sulphophenylazo)pyrazole-3-carboxylate) using Chinese hamster fibroblast cell line CHL. The cells were exposed to the test material at three different doses with 2.5 mg/plate being the highest dose for 24 and 48 hr. In the present studies, no metabolic activation systems were applied. The incidence of polyploid cells as well as of cells with structural chromosomal aberrations such as chromatid or chromosome gaps, breaks, exchanges, ring formations, fragmentations and others, was recorded on each culture plate. Untreated cells and solvent-treated cells served as negative controls, in which the incidence of aberrations was usually less than 3.0%. The results were considered to be negative if the incidence was less than 4.9%, equivocal if it was between 5.0 and 9.9%, and positive if it was more than 10.0%. The incidence of chromosome aberration in Chinese hamster fibroblast cell line for the test chemical tartrazine was considered to be more than 10% in the absence of metabolic activation system during the 48 hrs study duration and hence tartrazine is mutagenic in vitro.

In the same study by Ishidate et al, Gene mutation toxicity study was performed to determine the mutagenic nature of tartrazine (RA CAS no 1934 -21 -0). The study was performed using S. typhimurium strains TA92, TA1535, TA100, TA1537, TA94 and TA98 with and without S9 metabolic activation system. The test was performed as per the preincubation assay at six different concentrations with 5.0 mg/plate being the maximum concentration. The chemical was dissolved in distilled water. Preincubation was performed for 20 mins and the exposure duration was for 48 hrs. The result was considered positive if the number of colonies found was twice the number in the control (exposed to the appropriate solvent or untreated). Tartrazine did not induce a doubling of revertant colonies over the control using S. typhimurium strains TA92, TA1535, TA100, TA1537, TA94 and TA98 in the presence and absence of S9 metabolic activation system and hence the chemical is not likely to classify as a gene mutant in vitro.

Ivett et al (Environmental and Molecular Mutagenesis, 1989) performed chromosomal aberration study for another strcturally and functionally simlar read across chemical. Chromosomal aberration tests were carried out to determine the mutagenic nature of FD and C yellow 6 (RA CAS no 2783 -94 -0; IUPAC name: disodium (5E)-6-oxo-5-[(4-sulfonatophenyl)hydrazinylidene]naphthalene-2-sulfonate). The study was performed using Chinese hamster ovary cells in the presence and absence of S9 metabolic activation system. FD and C yellow no 6 was dissolved in medium and used at dose levels of up to 5 mg/mL. In the assays for chromosomal aberrations, the top dose (TD) was based on toxicity, solubility, or the upper testing limit (5 mg/ml). In the chromosome aberration assay without activation, cells were exposed to the test chemical for 8 hr. The test chemical was washed off, and the cells were treated with 0.1 µg/ml Colcemid for 2-2.5 hr. With metabolic activation, the cells were exposed to the test chemical plus the metabolic activation mixture for 2 hr, washed, incubated for 8 hr, and then treated with Colcemid for 2-2.5 hr. A delayed harvest was used in the aberration assay in most instances when cell cycle delay was observed in the SCE assay. In these tests the cell growth period was extended to about 20 hr. Air-dried slides were coded and stained with Giemsa. One hundred to 200 cells from each of the three highest scorable doses were analyzed. All aberrations were individually classified (e.g., chromatid breaks, chromosome breaks, triradials, etc.). These data were combined as the percent of cells with simple (deletions), complex (exchanges), and total (simple, complex and other) aberrations. FD and C yellow no 6 did not induce chromosome aberrations in Chinese hamster ovary cells (CHO) in the presence and absence of S9 metabolic activation system and hence it is not likely to classify as a gene mutant in vitro.

Gene mutation toxicity study was also performed by CHung et al (Applied and Environmental Microbiology, 1981) to determine the mutagenic nature of Sunset yellow (RA CAS no 2783 -94 -0, IUPAC name: disodium (5E)-6-oxo-5-[(4-sulfonatophenyl)hydrazinylidene]naphthalene-2-sulfonate). The study was performed by the standard plate incorporation assay using Salmonella typhimurium strains TA1535, TA1537, TA1538, TA98, and TA100 with and without S9 metabolic activation system. The test chemical was dissolved in DMSO and upto a maximum nontoxic dose of 5000 µg/plate. Concurrent solvent and positive controls were also included in the study. Sunset yellow did not induce gene mutation in Salmonella typhimurium strains TA1535, TA1537, TA1538, TA98, and TA100 in the presence and absence of S9 metabolic activation system and hence it is not likely to classify as a gene mutant in vitro.

Based on the data available for the target chemical and its read across, aluminum tris(4-{[3-carboxy-5-oxo-1-(4-sulfophenyl)-4,5-dihydro-1H-pyrazol-4-yl]diazenyl}benzenesulfonate) does not exhibit gene mutation in vitro. Hence it is not likely to classify as a gene mutant as per the criteria mentioned in CLP regulation. The read across chemical with CAS no 1934 -21 -0 is the parent compound for aluminum tris(4-{[3-carboxy-5-oxo-1-(4-sulfophenyl)-4,5-dihydro-1H-pyrazol-4-yl]diazenyl}benzenesulfonate) and its depicts the non mutagenic nature in the Ames test and the chromosomal aberration study performed with metabolic activation system. Considering this and the other read across data, the test chemical aluminum tris(4-{[3-carboxy-5-oxo-1-(4-sulfophenyl)-4,5-dihydro-1H-pyrazol-4-yl]diazenyl}benzenesulfonate) is considered to be non-mutagenic in vitro.

Justification for classification or non-classification

Based on the data available for the target chemical and its read across, aluminum tris(4-{[3-carboxy-5-oxo-1-(4-sulfophenyl)-4,5-dihydro-1H-pyrazol-4-yl] diazenyl}benzenesulfonate) (CAS no 12225 -21 -7) does not exhibit gene mutation in vitro. Hence it is not likely to classify as a gene mutant as per the criteria mentioned in CLP regulation.