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EC number: 293-766-2 | CAS number: 91082-52-9
- Life Cycle description
- Uses advised against
- Endpoint summary
- Appearance / physical state / colour
- Melting point / freezing point
- Boiling point
- Density
- Particle size distribution (Granulometry)
- Vapour pressure
- Partition coefficient
- Water solubility
- Solubility in organic solvents / fat solubility
- Surface tension
- Flash point
- Auto flammability
- Flammability
- Explosiveness
- Oxidising properties
- Oxidation reduction potential
- Stability in organic solvents and identity of relevant degradation products
- Storage stability and reactivity towards container material
- Stability: thermal, sunlight, metals
- pH
- Dissociation constant
- Viscosity
- Additional physico-chemical information
- Additional physico-chemical properties of nanomaterials
- Nanomaterial agglomeration / aggregation
- Nanomaterial crystalline phase
- Nanomaterial crystallite and grain size
- Nanomaterial aspect ratio / shape
- Nanomaterial specific surface area
- Nanomaterial Zeta potential
- Nanomaterial surface chemistry
- Nanomaterial dustiness
- Nanomaterial porosity
- Nanomaterial pour density
- Nanomaterial photocatalytic activity
- Nanomaterial radical formation potential
- Nanomaterial catalytic activity
- Endpoint summary
- Stability
- Biodegradation
- Bioaccumulation
- Transport and distribution
- Environmental data
- Additional information on environmental fate and behaviour
- Ecotoxicological Summary
- Aquatic toxicity
- Endpoint summary
- Short-term toxicity to fish
- Long-term toxicity to fish
- Short-term toxicity to aquatic invertebrates
- Long-term toxicity to aquatic invertebrates
- Toxicity to aquatic algae and cyanobacteria
- Toxicity to aquatic plants other than algae
- Toxicity to microorganisms
- Endocrine disrupter testing in aquatic vertebrates – in vivo
- Toxicity to other aquatic organisms
- Sediment toxicity
- Terrestrial toxicity
- Biological effects monitoring
- Biotransformation and kinetics
- Additional ecotoxological information
- Toxicological Summary
- Toxicokinetics, metabolism and distribution
- Acute Toxicity
- Irritation / corrosion
- Sensitisation
- Repeated dose toxicity
- Genetic toxicity
- Carcinogenicity
- Toxicity to reproduction
- Specific investigations
- Exposure related observations in humans
- Toxic effects on livestock and pets
- Additional toxicological data
Genetic toxicity: in vitro
Administrative data
- Endpoint:
- in vitro gene mutation study in bacteria
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Study period:
- Experimental start: 05 May 2016, Experimental completion: 20 May 2016; Final Report: 01 September 2016
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
Data source
Reference
- Reference Type:
- study report
- Title:
- Unnamed
- Year:
- 2 016
- Report date:
- 2016
Materials and methods
Test guideline
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 471 (Bacterial Reverse Mutation Assay)
- Version / remarks:
- 1997
- Deviations:
- no
- GLP compliance:
- yes (incl. QA statement)
- Type of assay:
- bacterial reverse mutation assay
Test material
- Reference substance name:
- Tar bases, coal, lutidine fraction
- EC Number:
- 293-766-2
- EC Name:
- Tar bases, coal, lutidine fraction
- Cas Number:
- 91082-52-9
- IUPAC Name:
- Tar bases, coal, lutidine fraction
- Test material form:
- liquid
Constituent 1
Method
Species / strain
- 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 prepared from male Sprague-Dawley derived rats
- Test concentrations with justification for top dose:
- 5, 15, 50, 150, 500, 1500 and 5000 µg/plate in the absence and presence of metabolic activation. The maximum concentration was selected based on the standard limit concentration recommended in the regulatory guidelines that the assay follows.
- Vehicle / solvent:
- - Vehicle(s)/solvent(s) used: DMSO, ACS reagent grade
- Justification for choice of solvent/vehicle: the test substances dissolved completely in the vehicle at the highest dose tested
Controls
- Negative solvent / vehicle controls:
- yes
- Positive controls:
- yes
- Positive control substance:
- 4-nitroquinoline-N-oxide
- 9-aminoacridine
- 2-nitrofluorene
- sodium azide
- benzo(a)pyrene
- other: 2-aminoanthracene
- Details on test system and experimental conditions:
- METHOD OF APPLICATION: in agar (plate incorporation); preincubation
- Cell density at seeding (if applicable): at least 10^9 per mL at test begin
DURATION
- Preincubation period: 30 minutes
- Exposure duration: 48 to 72 hours
- Expression time (cells in growth medium): 10 hours
DETERMINATION OF CYTOTOXICITY
- Method: relative total growth
- Any supplementary information relevant to cytotoxicity: toxicity was observed as a thin background lawn of non-revertant colonies and/or a reduction in the number of revertant colonies
NUMBER OF REPLICATIONS: 3 - Rationale for test conditions:
- Following relevant test guideline
- Evaluation criteria:
- Criteria for valid test:
- the mean of the vehicle control revertant colony numbers for each strain should lie within or close to the current historical control range for the laboratory (maintained as a rolling record over two years or a minimum of 20 data sets)
- the positive control compounds must induce an increase in mean revertant colony numbers of at least twice that of the concurrent vehicle control
- mean viable cell counts in the 10-hour bacterial cultures must be at least 10^9 per mL
- a minimum of five analysable concentrations must be present with at least four showing no signs of toxic effects, evident as bacterial inhibition and/or a reduction in the number of revertants below the indication factor of 0.5
If exposure to a test substance produces a reproducible increase in mean revertant colony numbers of at least twice that of the vehicle controls, with some evidence of a positive concentration-response relationship, it is considered to exhibit mutagenic effects.
If exposure to a test substance does not produce a reproducible increase in mean revertant colony numbers, it is considered to show no evidence of mutagenic activity.
If the results obtained fail to satisfy the criteria for a clear "positive" or "negative" response, even after additional testing, the test data may be subjected to analysis to determine the statistical significance of any increases in revertant colony numbers. In general, treatment-associated increases in mean revertant colony numbers below two or three times those of the vehicle controls are not considered biologically important. It should be noted that it is acceptable to conclude an equivocal response if no clear results can be obtained. - Statistics:
- The statistical procedures used are those described by Mahon et al (1989) and are usually Dunnett's test followed, if appropriate, by trend analysis. Biological importance will be considered along with statistical significance.
Results and discussion
Test resultsopen allclose all
- Species / strain:
- S. typhimurium TA 1535
- Metabolic activation:
- with and without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- no cytotoxicity nor precipitates, but tested up to recommended limit concentrations
- Vehicle controls validity:
- valid
- Positive controls validity:
- valid
- Species / strain:
- S. typhimurium TA 1537
- Metabolic activation:
- with and without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- no cytotoxicity nor precipitates, but tested up to recommended limit concentrations
- Vehicle controls validity:
- valid
- Positive controls validity:
- valid
- Species / strain:
- S. typhimurium TA 100
- Metabolic activation:
- with and without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- no cytotoxicity nor precipitates, but tested up to recommended limit concentrations
- Vehicle controls validity:
- valid
- Positive controls validity:
- valid
- Species / strain:
- S. typhimurium TA 98
- Metabolic activation:
- with and without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- cytotoxicity
- Vehicle controls validity:
- valid
- Positive controls validity:
- valid
- Species / strain:
- E. coli WP2 uvr A
- Metabolic activation:
- with and without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- no cytotoxicity nor precipitates, but tested up to recommended limit concentrations
- Vehicle controls validity:
- valid
- Positive controls validity:
- valid
- Additional information on results:
- TEST-SPECIFIC CONFOUNDING FACTORS
- Effects of pH: no fluctuations in pH of the medium were observed at 2000 μg/mL of more than 1.0 unit compared with the vehicle control
- Effects of osmolality: no fluctuations in osmolality of the medium of more than 50 mOsm/kg were observed compared with the vehicle control
- Evaporation from medium: not reported
- Precipitation: no precipitation was observed
RANGE-FINDING/SCREENING STUDIES:
HISTORICAL CONTROL DATA (with ranges, means and standard deviation and confidence interval (e.g. 95%)
- See under any other information on results
Any other information on results incl. tables
Table 1: Results of Experiment 1, plate incorporation
Experiment 1, plate incorporation, no metabolic activation |
||||||||||||||||||||||||
Strain |
Addition |
Concentration per plate [µg] |
Mean revertant number per plate |
Standard deviation |
Fold increase over vehicle |
Individual revertant counts |
||||||||||||||||||
TA98 |
DMSO |
|
31.0 |
1.7 |
|
32 |
29 |
32 |
||||||||||||||||
Test substance |
5 |
26.7 |
6.5 |
0.9 |
27 |
33 |
20 |
|||||||||||||||||
15 |
24.3 |
1.5 |
0.8 |
24 |
23 |
26 |
||||||||||||||||||
50 |
12.7 |
4.6 |
0.4 |
18 |
10 |
10 |
||||||||||||||||||
150 |
7.7 |
2.3 |
0.2 |
9 |
9 |
5 |
||||||||||||||||||
500 |
9.0 |
4.4 |
0.3 |
4 |
12 |
11 |
||||||||||||||||||
1500 |
12.7 |
2.1 |
0.4 |
15S |
11S |
12S |
||||||||||||||||||
5000 |
8.7 |
2.1 |
0.3 |
7S |
11S |
8S |
||||||||||||||||||
|
||||||||||||||||||||||||
TA100 |
DMSO |
|
141.0 |
6.9 |
|
137 |
149 |
137 |
||||||||||||||||
Test substance |
5 |
133.0 |
11.4 |
0.9 |
120 |
138 |
141 |
|||||||||||||||||
15 |
149.7 |
2.5 |
1.1 |
152 |
150 |
147 |
||||||||||||||||||
50 |
107.3 |
27.6 |
0.8 |
139 |
88 |
95 |
||||||||||||||||||
150 |
104.7 |
5.8 |
0.7 |
108 |
98 |
108 |
||||||||||||||||||
500 |
93.0 |
11.8 |
0.7 |
106 |
90 |
83 |
||||||||||||||||||
1500 |
114.7 |
4.0 |
0.8 |
117 |
110 |
117 |
||||||||||||||||||
5000 |
111.7 |
7.0 |
0.8 |
119 |
105 |
111 |
||||||||||||||||||
|
||||||||||||||||||||||||
TA1535 |
DMSO |
|
22.3 |
9.5 |
|
13 |
32 |
22 |
||||||||||||||||
Test substance |
5 |
21.0 |
1.0 |
0.9 |
21 |
22 |
20 |
|||||||||||||||||
15 |
18.0 |
3.5 |
0.8 |
16 |
16 |
22 |
||||||||||||||||||
50 |
18.0 |
8.0 |
0.8 |
26 |
10 |
18 |
||||||||||||||||||
150 |
14.0 |
6.1 |
0.6 |
11 |
10 |
21 |
||||||||||||||||||
500 |
18.7 |
4.7 |
0.8 |
17 |
15 |
24 |
||||||||||||||||||
1500 |
17.0 |
4.6 |
0.8 |
21 |
12 |
18 |
||||||||||||||||||
5000 |
21.7 |
7.8 |
1.0 |
24 |
28 |
13 |
||||||||||||||||||
|
||||||||||||||||||||||||
TA1537 |
DMSO |
|
17.7 |
4.5 |
|
13 |
18 |
22 |
||||||||||||||||
Test substance |
5 |
16.7 |
7.1 |
0.9 |
18 |
9 |
23 |
|||||||||||||||||
15 |
19.0 |
5.6 |
1.1 |
24 |
13 |
20 |
||||||||||||||||||
50 |
12.3 |
3.1 |
0.7 |
15 |
9 |
13 |
||||||||||||||||||
150 |
12.3 |
3.2 |
0.7 |
11 |
10 |
16 |
||||||||||||||||||
500 |
12.7 |
2.1 |
0.7 |
12 |
15 |
11 |
||||||||||||||||||
1500 |
11.7 |
1.5 |
0.7 |
10 |
13 |
12 |
||||||||||||||||||
5000 |
17.7 |
9.0 |
1.0 |
13 |
28 |
12 |
||||||||||||||||||
|
||||||||||||||||||||||||
WP2 uvrA (pKM101) |
DMSO |
|
156.7 |
17.7 |
|
177 |
148 |
145 |
||||||||||||||||
Test substance |
5 |
161.3 |
9.7 |
1.0 |
153 |
159 |
172 |
|||||||||||||||||
15 |
164.0 |
5.6 |
1.0 |
169 |
165 |
158 |
||||||||||||||||||
50 |
128.0 |
13.0 |
0.8 |
143 |
120 |
121 |
||||||||||||||||||
150 |
121.7 |
11.2 |
0.8 |
109 |
130 |
126 |
||||||||||||||||||
500 |
125.7 |
9.1 |
0.8 |
116 |
127 |
134 |
||||||||||||||||||
1500 |
162.0 |
10.6 |
1.0 |
174 |
158 |
154 |
||||||||||||||||||
5000 |
124.3 |
16.3 |
0.8 |
142 |
110 |
121 |
||||||||||||||||||
S: slight thinning of background lawn |
||||||||||||||||||||||||
Experiment 1, plate incorporation, no metabolic activation, positive controls |
||||||||||||||||||||||||
TA98 |
2-nitrofluorene |
2 |
243.3 |
9.2 |
7.8 |
238 |
238 |
254 |
||||||||||||||||
TA100 |
Sodium azide |
2 |
540.0 |
6.2 |
3.8 |
542 |
533 |
545 |
||||||||||||||||
TA1535 |
Sodium azide |
2 |
828.3 |
44.5 |
37.1 |
821 |
876 |
788 |
||||||||||||||||
TA1537 |
9-aminoacridine |
50 |
202.0 |
19.5 |
11.4 |
182 |
221 |
203 |
||||||||||||||||
WP2 uvrA |
4-nitroquinoline-1-oxide |
2 |
847.7 |
95.1 |
5.4 |
740 |
920 |
883 |
||||||||||||||||
Experiment 1, plate incorporation, viability |
||||||||||||||||||||||||
|
|
Strain |
|
Mean counts per plate |
Standard deviation |
Individual counts (100 µL aliquots of 10^-6 dilution of 10-hour culture) |
||||||||||||||||||
|
|
TA98 |
Viability |
244.7 |
17.0 |
225 |
255 |
254 |
||||||||||||||||
|
|
TA100 |
Viability |
327.0 |
12.3 |
332 |
313 |
336 |
||||||||||||||||
|
|
TA1535 |
Viability |
351.0 |
89.7 |
331 |
449 |
273 |
||||||||||||||||
|
|
TA1537 |
Viability |
242.7 |
9.1 |
251 |
233 |
244 |
||||||||||||||||
|
|
WP2 uvrA |
Viability |
330.0 |
13.2 |
345 |
320 |
324 |
||||||||||||||||
Experiment 1, plate incorporation, with metabolic activation |
||||||||||||||||||||||||
Strain |
Addition |
Concentration per plate [µg] |
Mean revertant number per plate |
Standard deviation |
Fold increase over vehicle |
Individual revertant counts |
||||||||||||||||||
TA98 |
DMSO |
|
31.7 |
6.4 |
|
39 |
27 |
29 |
||||||||||||||||
Test substance |
5 |
20.0 |
3.6 |
0.6 |
23 |
21 |
16 |
|||||||||||||||||
15 |
19.3 |
3.1 |
0.6 |
22 |
20 |
16 |
||||||||||||||||||
50 |
20.3 |
3.8 |
0.6 |
16 |
23 |
22 |
||||||||||||||||||
150 |
21.3 |
0.6 |
0.7 |
22 |
21 |
21 |
||||||||||||||||||
500 |
26.3 |
2.1 |
0.8 |
27 |
28 |
24 |
||||||||||||||||||
1500 |
21.7 |
2.5 |
0.7 |
24S |
22S |
19S |
||||||||||||||||||
5000 |
22.7 |
3.8 |
0.7 |
21S |
20S |
27S |
||||||||||||||||||
|
||||||||||||||||||||||||
TA100 |
DMSO |
|
150.0 |
14.9 |
|
133 |
161 |
156 |
||||||||||||||||
Test substance |
5 |
109.3 |
9.2 |
0.7 |
104 |
120 |
104 |
|||||||||||||||||
15 |
108.0 |
8.0 |
0.7 |
116 |
108 |
100 |
||||||||||||||||||
50 |
110.0 |
6.6 |
0.7 |
111 |
116 |
103 |
||||||||||||||||||
150 |
108.0 |
15.9 |
0.7 |
114 |
90 |
120 |
||||||||||||||||||
500 |
126.3 |
10.2 |
0.8 |
138 |
119 |
122 |
||||||||||||||||||
1500 |
131.3 |
5.0 |
0.9 |
136 |
126 |
132 |
||||||||||||||||||
5000 |
134.7 |
9.0 |
0.9 |
134 |
126 |
144 |
||||||||||||||||||
|
||||||||||||||||||||||||
TA1535 |
DMSO |
|
14.0 |
2.6 |
|
15 |
16 |
11 |
||||||||||||||||
Test substance |
5 |
14.0 |
3.5 |
1.0 |
12 |
12 |
18 |
|||||||||||||||||
15 |
10.7 |
4.5 |
0.8 |
11 |
15 |
6 |
||||||||||||||||||
50 |
14.7 |
7.5 |
1.0 |
7 |
22 |
15 |
||||||||||||||||||
150 |
9.3 |
3.8 |
0.7 |
12 |
11 |
5 |
||||||||||||||||||
500 |
16.7 |
4.5 |
1.2 |
12 |
21 |
17 |
||||||||||||||||||
1500 |
20.3 |
2.9 |
1.5 |
22 |
17 |
22 |
||||||||||||||||||
5000 |
13.3 |
8.1 |
1.0 |
12 |
22 |
6 |
||||||||||||||||||
|
||||||||||||||||||||||||
TA1537 |
DMSO |
|
19.7 |
6.7 |
|
23 |
24 |
12 |
||||||||||||||||
Test substance |
5 |
17.0 |
6.9 |
0.9 |
21 |
21 |
9 |
|||||||||||||||||
15 |
14.7 |
1.5 |
0.7 |
13 |
15 |
16 |
||||||||||||||||||
50 |
15.0 |
4.4 |
0.8 |
18 |
10 |
17 |
||||||||||||||||||
150 |
15.7 |
6.4 |
0.8 |
23 |
12 |
12 |
||||||||||||||||||
500 |
14.3 |
3.8 |
0.7 |
10 |
16 |
17 |
||||||||||||||||||
1500 |
18.0 |
5.0 |
0.9 |
13 |
23 |
18 |
||||||||||||||||||
5000 |
17.0 |
5.3 |
0.9 |
13 |
15 |
23 |
||||||||||||||||||
|
||||||||||||||||||||||||
WP2 uvrA (pKM101) |
DMSO |
|
199.0 |
14.0 |
|
189 |
215 |
193 |
||||||||||||||||
Test substance |
5 |
152.3 |
4.7 |
0.8 |
154 |
147 |
156 |
|||||||||||||||||
15 |
167.7 |
17.8 |
0.8 |
160 |
155 |
188 |
||||||||||||||||||
50 |
184.0 |
19.0 |
0.9 |
189 |
200 |
163 |
||||||||||||||||||
150 |
174.3 |
3.8 |
0.9 |
176 |
170 |
177 |
||||||||||||||||||
500 |
182.3 |
5.9 |
0.9 |
178 |
180 |
189 |
||||||||||||||||||
1500 |
199.7 |
20.6 |
1.0 |
178 |
202 |
219 |
||||||||||||||||||
5000 |
152.3 |
22.3 |
0.8 |
178 |
138 |
141 |
||||||||||||||||||
S: slight thinning of background lawn |
||||||||||||||||||||||||
Experiment 1, plate incorporation, with metabolic activation, positive control |
||||||||||||||||||||||||
TA98 |
Benzo[a]pyrene |
5 |
139.0 |
7.2 |
4.4 |
141 |
131 |
145 |
||||||||||||||||
TA100 |
2-aminoanthracene |
5 |
817.7 |
388.6 |
5.5 |
1266 |
577 |
610 |
||||||||||||||||
TA1535 |
2-aminoanthracene |
5 |
542.0 |
50.9 |
38.7 |
488 |
589 |
549 |
||||||||||||||||
TA1537 |
Benzo[a]pyrene |
5 |
737 |
2.9 |
3.7 |
77 |
72 |
72 |
||||||||||||||||
WP2 uvrA |
2-aminoanthracene |
10 |
834.0 |
35.0 |
4.2 |
874 |
819 |
809 |
||||||||||||||||
Additional experiment 1, plate incorporation, no metabolic activation |
||||||||||||||||||||||||
Strain |
Addition |
Concentration per plate [µg] |
Mean revertant number per plate |
Standard deviation |
Fold increase over vehicle |
Individual revertant counts |
||||||||||||||||||
TA98 |
DMSO |
|
21.7 |
0.6 |
|
22 |
21 |
22 |
||||||||||||||||
Test substance |
0.5 |
17.3 |
0.6 |
0.8 |
17 |
17 |
18 |
|||||||||||||||||
1.0 |
17.3 |
3.2 |
0.8 |
15 |
16 |
21 |
||||||||||||||||||
5 |
12.0 |
1.7 |
0.6 |
10 |
13 |
13 |
||||||||||||||||||
15 |
19.3 |
3.1 |
0.9 |
22 |
16 |
20 |
||||||||||||||||||
50 |
21.0 |
0.0 |
1.0 |
21 |
21 |
21 |
||||||||||||||||||
150 |
20.0 |
7.0 |
0.9 |
17 |
28 |
15 |
||||||||||||||||||
500 |
13.3 |
3.5 |
0.6 |
13 |
10 |
17 |
||||||||||||||||||
1500 |
8.3 |
0.6 |
0.4 |
8S |
9S |
8S |
||||||||||||||||||
5000 |
6.0 |
1.0 |
0.3 |
6S |
7S |
5S |
||||||||||||||||||
S: slight thinning of background lawn |
||||||||||||||||||||||||
Additional experiment 1, plate incorporation, no metabolic activation, positive control |
||||||||||||||||||||||||
TA98 |
2-nitrofluorene |
2 |
262.7 |
16.9 |
12.1 |
255 |
282 |
251 |
||||||||||||||||
Additional experiment 1, plate incorporation, no metabolic activation, viability |
||||||||||||||||||||||||
Strain |
|
Mean counts per plate |
Standard deviation |
Individual counts (100 µL aliquots of 10^-6 dilution of 10-hour culture) |
||||||||||||||||||||
TA98 |
Viability |
211.0 |
29.5 |
177 |
226 |
230 |
Table 2: Results of Experiment 2, pre-incubation
Experiment 2, pre-incubation, no metabolic activation |
||||||||
Strain |
Addition |
Concentration per plate [µg] |
Mean revertant number per plate |
Standard deviation |
Fold increase over vehicle |
Individual revertant counts |
||
TA98 |
DMSO |
|
50.3 |
12.7 |
|
65 |
43 |
43 |
Test substance |
0.5 |
39.7 |
2.9 |
0.8 |
38 |
38 |
43 |
|
1.5 |
41.0 |
3.5 |
0.8 |
45 |
39 |
39 |
||
5 |
45.7 |
9.3 |
0.9 |
38 |
43 |
56 |
||
15 |
52.0 |
8.9 |
1.0 |
45 |
62 |
49 |
||
50 |
43.0 |
0.0 |
0.9 |
53 |
43 |
43 |
||
150 |
33.3 |
5.5 |
0.7 |
39 |
28 |
33 |
||
500 |
19.3 |
1.2 |
0.4 |
20S |
15S |
20S |
||
1500 |
19.7 |
1.5 |
0.4 |
21T |
20T |
18T |
||
5000 |
20.3 |
3.1 |
0.4 |
17T |
23T |
21T |
||
|
||||||||
TA100 |
DMSO |
|
146.0 |
9.5 |
|
137 |
156 |
145 |
Test substance |
5 |
152.7 |
5.5 |
1.0 |
158 |
147 |
153 |
|
15 |
151.7 |
23.4 |
1.0 |
169 |
161 |
125 |
||
50 |
109.7 |
26.1 |
0.8 |
139 |
89 |
101 |
||
150 |
109.0 |
10.6 |
0.7 |
101 |
121 |
105 |
||
500 |
107.0 |
5.6 |
0.7 |
108 |
101 |
112 |
||
1500 |
111.7 |
13.5 |
0.8 |
98 |
112 |
125 |
||
5000 |
121.7 |
24.6 |
0.8 |
94 |
141 |
130 |
||
|
||||||||
TA1535 |
DMSO |
|
16.3 |
5.7 |
|
21 |
18 |
10 |
Test substance |
5 |
13.3 |
2.5 |
0.8 |
11 |
13 |
16 |
|
15 |
20.7 |
12.9 |
1.3 |
10 |
17 |
35 |
||
50 |
14.3 |
5.8 |
0.9 |
21 |
11 |
11 |
||
150 |
12.0 |
1.7 |
0.7 |
13 |
10 |
13 |
||
500 |
13.3 |
2.3 |
0.8 |
16 |
12 |
12 |
||
1500 |
18.3 |
2.5 |
1.1 |
21 |
16 |
18 |
||
5000 |
15.3 |
5.8 |
0.9 |
22 |
12 |
12 |
||
|
||||||||
TA1537 |
DMSO |
|
10.0 |
2.6 |
|
11 |
12 |
7 |
Test substance |
5 |
13.7 |
8.5 |
1.4 |
17 |
4 |
20 |
|
15 |
11.0 |
6.2 |
1.1 |
6 |
18 |
9 |
||
50 |
6.0 |
3.5 |
0.6 |
4 |
10 |
4 |
||
150 |
11.7 |
1.2 |
1.2 |
11 |
13 |
11 |
||
500 |
6.0 |
3.6 |
0.6 |
2 |
7 |
9 |
||
1500 |
6.7 |
2.5 |
0.7 |
4 |
7 |
9 |
||
5000 |
5.7 |
2.9 |
0.6 |
4 |
9 |
4 |
||
|
||||||||
WP2 uvrA (pKM101) |
DMSO |
|
163.7 |
24.0 |
|
177 |
136 |
178 |
Test substance |
5 |
168.0 |
6.6 |
1.0 |
161 |
174 |
169 |
|
15 |
193.7 |
25.0 |
1.2 |
172 |
221 |
188 |
||
50 |
135.0 |
17.4 |
0.8 |
155 |
123 |
127 |
||
150 |
137.7 |
6.7 |
0.8 |
142 |
130 |
141 |
||
500 |
138.3 |
10.2 |
0.8 |
134 |
131 |
150 |
||
1500 |
136.0 |
3.0 |
0.8 |
136 |
139 |
133 |
||
5000 |
103.7 |
9.2 |
0.6 |
93 |
109 |
109 |
||
Experiment 2, pre-incubation, no metabolic activation, positive control |
||||||||
TA98 |
2-nitrofluorene |
2 |
237.2 |
42.9 |
4.7 |
189 |
271 |
252 |
TA100 |
Sodium azide |
2 |
652.7 |
31.8 |
4.5 |
630 |
689 |
639 |
TA1535 |
Sodium azide |
2 |
672.7 |
58.5 |
41.2 |
732 |
671 |
615 |
TA1537 |
9-aminoacridine |
50 |
191.3 |
28.9 |
28.9 |
209 |
207 |
158 |
WP2 uvrA |
4-nitroquinoline-1-oxide |
2 |
1028.7 |
12.1 |
6.3 |
1038 |
1033 |
1015 |
S: slight thinning of background lawn; T: thinning of background lawn |
||||||||
Experiment 2, pre-incubation, with metabolic activation |
||||||||
Strain |
Addition |
Concentration per plate [µg] |
Mean revertant number per plate |
Standard deviation |
Fold increase over vehicle |
Individual revertant counts |
||
TA98 |
DMSO |
|
30.0 |
1.7 |
|
32 |
29 |
29 |
Test substance |
0.5 |
24.3 |
7.5 |
0.8 |
32 |
17 |
24 |
|
1.5 |
25.0 |
5.2 |
0.8 |
22 |
22 |
31 |
||
5 |
32.0 |
1.7 |
1.1 |
31 |
31 |
34 |
||
15 |
33.0 |
3.5 |
1.1 |
31 |
37 |
31 |
||
50 |
21.3 |
5.8 |
0.7 |
18 |
18 |
28 |
||
150 |
21.0 |
0.0 |
1.7 |
21 |
21 |
21 |
||
500 |
28.3 |
3.2 |
0.9 |
27S |
32S |
26S |
||
1500 |
24.0 |
2.0 |
0.8 |
26T |
22T |
24T |
||
5000 |
18.0 |
2.6 |
0.6 |
16T |
17T |
21T |
||
|
||||||||
TA100 |
DMSO |
|
142.0 |
10.0 |
|
142 |
132 |
152 |
Test substance |
5 |
100.7 |
7.6 |
0.7 |
104 |
92 |
106 |
|
15 |
110.0 |
4.6 |
0.8 |
106 |
109 |
115 |
||
50 |
114.3 |
11.6 |
0.8 |
101 |
122 |
120 |
||
150 |
124.3 |
13.1 |
0.9 |
138 |
112 |
123 |
||
500 |
132.3 |
13.3 |
0.9 |
117 |
141 |
139 |
||
1500 |
141.7 |
2.5 |
1.0 |
139 |
142 |
144 |
||
5000 |
145.0 |
12.8 |
1.0 |
148 |
156 |
131 |
||
|
||||||||
TA1535 |
DMSO |
|
15.0 |
3.0 |
|
12 |
15 |
18 |
Test substance |
5 |
10.7 |
4.7 |
0.7 |
16 |
9 |
7 |
|
15 |
13.7 |
1.2 |
0.9 |
15 |
13 |
13 |
||
50 |
11.0 |
6.2 |
0.7 |
6 |
9 |
18 |
||
150 |
12.3 |
7.1 |
0.8 |
6 |
11 |
20 |
||
500 |
16.0 |
8.8 |
1.1 |
10 |
26 |
12 |
||
1500 |
10.7 |
8.1 |
0.7 |
18 |
2 |
12 |
||
5000 |
11.0 |
1.7 |
0.7 |
10 |
13 |
10 |
||
|
||||||||
TA1537 |
DMSO |
|
16.7 |
4.5 |
|
21 |
12 |
17 |
Test substance |
5 |
10.3 |
1.5 |
0.6 |
12 |
9 |
10 |
|
15 |
10.7 |
4.5 |
0.6 |
15 |
11 |
6 |
||
50 |
12.0 |
0.0 |
0.7 |
12 |
12 |
12 |
||
150 |
13.7 |
2.3 |
0.8 |
15 |
15 |
11 |
||
500 |
16.3 |
3.5 |
1.0 |
13 |
20 |
16 |
||
1500 |
12.0 |
1.7 |
0.7 |
13 |
10 |
13 |
||
5000 |
12.7 |
3.8 |
0.8 |
11 |
10 |
17 |
||
|
||||||||
WP2 uvrA (pKM101) |
DMSO |
|
200.7 |
15.0 |
|
215 |
185 |
202 |
Test substance |
5 |
177.0 |
39.1 |
0.9 |
132 |
202 |
197 |
|
15 |
173.3 |
15.4 |
0.9 |
166 |
191 |
163 |
||
50 |
166.0 |
10.5 |
0.8 |
156 |
177 |
165 |
||
150 |
176.3 |
11.7 |
0.9 |
163 |
185 |
181 |
||
500 |
164.7 |
5.5 |
0.8 |
165 |
159 |
170 |
||
1500 |
157.7 |
7.4 |
0.8 |
155 |
166 |
152 |
||
5000 |
143.0 |
5.3 |
0.7 |
139 |
149 |
141 |
||
S: slight thinning of background lawn; T: thinning of background lawn |
||||||||
Experiment 2, pre-incubation, with metabolic activation, positive control |
||||||||
TA98 |
Benzo[a]pyrene |
5 |
175.7 |
1.2 |
5.9 |
175 |
177 |
175 |
TA100 |
2-aminoanthracene |
5 |
2141.7 |
377.8 |
15.1 |
1711 |
2297 |
2417 |
TA1535 |
2-aminoanthracene |
5 |
361.0 |
19.5 |
24.1 |
346 |
354 |
383 |
TA1537 |
Benzo[a]pyrene |
5 |
159.3 |
1.2 |
9.6 |
158 |
160 |
160 |
WP2 uvrA |
2-aminoanthracene |
10 |
1038.0 |
71.5 |
5.2 |
1109 |
1039 |
966 |
Experiment 2, pre-incubation, viability |
||||||||
|
|
Strain |
|
Mean counts per plate |
Standard deviation |
Individual colony counts (100 µL aliquots of 10^-6 dilution of 10-hour culture) |
||
|
|
TA98 |
Viability |
150.0 |
9.8 |
142 |
147 |
161 |
|
|
TA100 |
Viability |
138.3 |
8.4 |
148 |
133 |
134 |
|
|
TA1535 |
Viability |
136.3 |
9.3 |
132 |
130 |
147 |
|
|
TA1537 |
Viability |
111.7 |
21.1 |
99 |
100 |
136 |
|
|
WP2 uvrA (pKM101) |
Viability |
188.7 |
23.1 |
167 |
213 |
186 |
Table 3: Historical control data
DMSO |
||||||||||
|
TA100 |
TA1535 |
WP2 uvrA (pKM101) |
TA98 |
TA1537 |
|||||
S9Mix |
- |
+ |
- |
+ |
- |
+ |
- |
+ |
- |
+ |
Max |
234 |
237 |
47 |
55 |
221 |
280 |
78 |
84 |
63 |
50 |
Min |
103 |
91 |
11 |
11 |
54 |
56 |
24 |
27 |
8 |
15 |
Mean |
158 |
168 |
26 |
23 |
193 |
193 |
42 |
55 |
21 |
32 |
No. of values |
147 |
145 |
147 |
141 |
131 |
131 |
153 |
149 |
146 |
143 |
St.dev. |
24 |
27 |
6 |
7 |
37 |
37 |
9 |
12 |
7 |
7 |
Upper 95% limit |
206 |
222 |
38 |
36 |
266 |
266 |
59 |
80 |
35 |
46 |
Lower 95% limit |
110 |
114 |
14 |
10 |
121 |
121 |
24 |
31 |
6 |
18 |
Positive controls |
||||||||||
|
TA100 |
TA1535 |
WP2 uvrA (pKM101) |
TA98 |
TA1537 |
|||||
|
NaN3 |
AAN |
NaN3 |
AAN |
NQO |
AAN |
2NF |
B[a]P |
AAC |
B[a]P |
S9Mix |
- |
+ |
- |
+ |
- |
+ |
- |
+ |
- |
+ |
Conc. (µg/plate) |
2 |
5 |
2 |
5 |
2 |
10 |
2 |
5 |
50 |
5 |
Max |
2776 |
4210 |
1255 |
716 |
3730 |
1923 |
802 |
648 |
2181 |
609 |
Min |
396 |
425 |
209 |
147 |
639 |
352 |
82 |
90 |
87 |
84 |
Mean |
1017 |
2017 |
799 |
378 |
2192 |
974 |
246 |
270 |
370 |
159 |
No. of values |
198 |
196 |
199 |
194 |
189 |
186 |
205 |
202 |
196 |
193 |
St.dev. |
292 |
763 |
219 |
127 |
651 |
368 |
123 |
94 |
270 |
58 |
NaN3: Sodium azide; 2NF: 2-nitrofluorene; AAC: 9-aminoacridine; B[a]P: Benzo[a]pyrene; AAN: 2-aminoanthracene |
Applicant's summary and conclusion
- Conclusions:
- The test substance showed no evidence of mutagenic activity in the tested strains of S. typhimurium and E. coli in the absence or presence of metabolic activation.
- Executive summary:
The mutagenic potential of the substance was studied in an in vitro bacterial reverse mutation (Ames) study in accordance with OECD TG 471 (1997) under GLP. The experiment is considered relevant, adequate and conclusive.
Experiments were conducted with the four histidine-dependent auxotrophic mutants of Salmonella typhimurium strains TA98, TA100, TA1535 and TA1537, and in addition with the tryptophan dependent mutant of Escherichia coli strain WP2 uvrA (pKM101) that were exposed to the test substance dissolved in dimethyl sulfoxide (DMSO).
Two independent mutation experiments were performed in the presence and absence of liver preparations (S9 mix) from rats treated with phenobarbital and 5,6-benzoflavone. The first experiment was a standard plate incorporation assay; the second experiment included a pre-incubation stage. Concentrations of up to 5000 µg/plate were tested, which is the standard limit concentration recommended in the current test guideline. In the first experiment, toxicity occurred, observed as a thin background lawn of non-revertant colonies and/or a reduction in the number of revertant colonies, following exposure to the test substance in strain TA98 at 50 µg/plate and above in the absence of S9 mix and at 1500 µg/plate and above in the presence of S9 mix. Strain TA98, in the absence of S9 mix, did not fulfil the criteria of a valid experiment and an additional plain incorporation experiment was therefore with this strain. In the second experiment, a pre-incubation test, toxicity (observed as thinning of the background lawn of non-revertant colonies, together with a reduction in revertant colony numbers) was obtained in strain TA98 following exposure to 500 µg/plate and above in the absence and presence of S9 mix. No precipitate was observed on plates following exposure to the test substance in both experiments. The concurrent positive controls verified the sensitivity of the assay and the metabolising activity of the S9 mix. The mean revertant colony numbers for the vehicle controls were within or close to the historical control range for the lab. The concurrent sterility controls demonstrated the absence of microbial contamination of the S9 mix, buffer or test substance formulation. No evidence of mutagenic activity of the test substance was observed at any tested concentration in either experiments in the absence or presence of S9 mix, and it was concluded that the test substance was not mutagenic in this Ames test.
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