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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:
- 2019-12-03 to 2020-01-16
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
Data source
Reference
- Reference Type:
- study report
- Title:
- Unnamed
- Year:
- 2 020
- Report date:
- 2020
Materials and methods
Test guideline
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 471 (Bacterial Reverse Mutation Assay)
- Version / remarks:
- adopted 21 July, 1997
- Deviations:
- no
- GLP compliance:
- yes (incl. QA statement)
- Type of assay:
- bacterial reverse mutation assay
Test material
- Reference substance name:
- Bis(glycinato)copper
- EC Number:
- 236-783-2
- EC Name:
- Bis(glycinato)copper
- Cas Number:
- 13479-54-4
- Molecular formula:
- C4H8CuN2O4
- IUPAC Name:
- copper;2-aminoacetate
- Test material form:
- solid: granular
- Details on test material:
- Appearance Grey to bluish powder/granule
Homogeneity homogeneous
Storage room temperature (20 ± 5 °C)
Stability stable under storage conditions
CAS No. 13479-54-4
EC-No. 236-783-2
Chemical Class organometallic chelate
Constituent 1
Method
- Target gene:
- his locus
Species / strain
- Species / strain / cell type:
- S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and TA 102
- Metabolic activation:
- with and without
- Metabolic activation system:
- Type and composition of metabolic activation system:
S9 liver Mix
- source of S9 : S9 was obtained by Trinova Biochem GmbH, Gießen.
- method of preparation of S9 mix: produced from the livers of male Sprague-Dawley rats which were treated with 500 mg Aroclor 1254/kg body weight intra-peritoneally.
- concentration or volume of S9 mix and S9 in the final culture medium: 500µL - Test concentrations with justification for top dose:
- nominal concentrations: 0, 50, 150, 500, 1500, 5000 µg/plate Experiment 1 and 0, 78, 156, 313, 625, 1250, 2500, 5000 /plate Experiment 2
- Vehicle / solvent:
- - Vehicle(s)/solvent(s) used: DMSO; demineralized water
- Justification for choice of solvent/vehicle: In a non-GLP pre-test, the solubility of the test item was tested in a concentration of 50 g/L in demineralized water, dimethyl sulfoxide (DMSO), acetone
The solid test item is not sufficiently soluble in any of the solvents. Based on the non-GLP pre-test, a test item suspension in demin. water was used, because this solvent shows the most consistent suspension with the test item and does not have any effects on the viability of the bacteria or the number of spontaneous revertants in the tested concentrations. The following nominal concentrations were prepared for the experiments 1b and 1d: 5000 µg/plate, 1500 µg/plate, 500 µg/plate, 150 µg/plate and 50 µg/plate. The following suspensions were stirred during the experiment: 5000 µg/plate, 1500 µg/plate, 500 µg/plate. The lower concentrations were dissolved completely and stirring was not necessary anymore. On the day of the start of the second experiment (2b), a test item stock solution containing 50 ± 5 g/L suspended in demin. water was prepared.
The following nominal concentrations were prepared for the second experiment: 5000 µg/plate, 2500 µg/plate, 1250 µg/plate, 625 µg/plate, 313 µg/plate, 156 µg/plate and 78 µg/plate. The suspensions were stirred during the experiment, except from 156 µg/plate and 78 µg/plate, they were dissolved completely and stirring was not necessary anymore.
DMSO was chosen due to the solubility of the positive controls 4-Nitro-1,2-phenylene diamine, benzo-a-pyrene and 2-amino-anthracene.
Controls
- Untreated negative controls:
- no
- Negative solvent / vehicle controls:
- yes
- True negative controls:
- no
- Positive controls:
- yes
- Positive control substance:
- sodium azide
- benzo(a)pyrene
- other: 4-Nitro-1,2-phenylene diamine: without metabolic activation, TA98, TA102, TA1537, 20 µg (TA98) and 30 µg (TA102 and TA1537) 2-Amino-anthracene: with metabolic activation, TA 100, TA102, TA1535, TA1537, 1µg (TA100, TA1535) and 2.4 µg (TA102, TA1537)
- Details on test system and experimental conditions:
- NUMBER OF REPLICATIONS:
- Number of cultures per concentration: triplicate for each with and without metabolic acitvation
- Number of independent experiments : 2
METHOD OF TREATMENT/ EXPOSURE:
- Cell density at seeding (if applicable): > E+09 cells/mL
- Test substance added in agar (plate incorporation)and in the second experiment with preincubation
METHODS FOR MEASUREMENT OF CYTOTOXICITY
- Method: background growth inhibition
- Any supplementary information relevant to cytotoxicity: No cytotoxicity nor precipitation occurred with the test item during the experimental time.
- Rationale for test conditions:
- as recommended by OECD Testguideline 471
- Evaluation criteria:
- The colonies were counted visually and the numbers were recorded. A substance is considered to be mutagenic, if a reproducible increase with or without metabolic activation of revertant colonies per plate exceeding an increase factor of 2 for the bacteria strains TA98, TA100, TA102, TA1535 and TA1537 compared to vehicle controls in at least one strain can be observed. A concentration-related increase over the range tested is also taken as a sign of mutagenic activity. A substance is not mutagenic if it does not meet these criteria. If the criteria listed above are not clearly met, the results will be assessed as equivocal and will be discussed.
- Statistics:
- The colonies were counted visually and the numbers were recorded. A validated spread-sheet software (Microsoft Excel®) was used to calculate mean values and standard deviations of each treatment, solvent control and positive control.
Results and discussion
Test resultsopen allclose all
- Key result
- 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
- Untreated negative controls validity:
- not examined
- True negative controls validity:
- not examined
- Positive controls validity:
- other: The positive control of strain TA1537 was slightly outside the historical data range. However, since the number of revertants was increased this was not considered to reduce the validity of the test result.
- Key result
- 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
- Untreated negative controls validity:
- not examined
- True negative controls validity:
- not examined
- Positive controls validity:
- other: The positive control of strain TA1537 was slightly outside the historical data range. However, since the number of revertants was increased this was not considered to reduce the validity of the test result.
- Key result
- Species / strain:
- S. typhimurium TA 102
- 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
- Untreated negative controls validity:
- not examined
- True negative controls validity:
- not examined
- Positive controls validity:
- other: The positive control of strain TA1537 was slightly outside the historical data range. However, since the number of revertants was increased this was not considered to reduce the validity of the test result.
- Key result
- Species / strain:
- S. typhimurium TA 98
- 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
- Untreated negative controls validity:
- not examined
- True negative controls validity:
- not examined
- Positive controls validity:
- other:
- Remarks:
- The positive control of strain TA1537 was slightly outside the historical data range. However, since the number of revertants was increased this was not considered to reduce the validity of the test result.
- Key result
- 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
- Untreated negative controls validity:
- not examined
- True negative controls validity:
- not examined
- Positive controls validity:
- other: The positive control of strain TA1537 was slightly outside the historical data range. However, since the number of revertants was increased this was not considered to reduce the validity of the test result.
- Additional information on results:
- TEST-SPECIFIC CONFOUNDING FACTORS
- Data on pH: not reported
- Water solubility: Not soluble in demineralized water, suspensions were used in the experiments.
RANGE-FINDING/SCREENING STUDIES (if applicable):
In a non-GLP pre-test, the solubility of the test item was tested in a concentration of 50 g/L in demineralized water, dimethyl sulfoxide (DMSO), acetone.
The solid test item is not sufficiently soluble in any of the solvents. Based on the non-GLP pre-test, a test item suspension in demin. water was used, because this solvent shows the most consistent suspension with the test item and does not have any effects on the viability of the bacteria or the number of spontaneous revertants in the tested concentrations. The following nominal concentrations were prepared for the experiments 1b and 1d: 5000 µg/plate, 1500 µg/plate, 500 µg/plate, 150 µg/plate and 50 µg/plate. The following suspensions were stirred during the experiment: 5000 µg/plate, 1500 µg/plate, 500 µg/plate. The lower concentrations were dissolved completely and stirring was not necessary anymore. On the day of the start of the second experiment (2b), a test item stock solution containing 50 ± 5 g/L suspended in demin. water was prepared.
The following nominal concentrations were prepared for the second experiment: 5000 µg/plate, 2500 µg/plate, 1250 µg/plate, 625 µg/plate, 313 µg/plate, 156 µg/plate and 78 µg/plate. The suspensions were stirred during the experiment, except from 156 µg/plate and 78 µg/plate, they were dissolved completely and stirring was not necessary anymore.no soluble in demineralized water
STUDY RESULTS
- Concurrent vehicle negative and positive control data Please refer to 'Any other infomation on results incl. tables'
Ames test:
- Signs of toxicity : No
- Individual plate counts : Please refer to 'Any other information on results incl. tables'.
- Mean number of revertant colonies per plate and standard deviation: Please refer to 'Any other information on results incl. tables'.
HISTORICAL CONTROL DATA (with ranges, means and standard deviation, and 95% control limits for the distribution as well as the number of data)
- Positive historical control data: Please refer to 'Any other information on results incl. tables'.
- Negative (solvent/vehicle) historical control data: Please refer to 'Any other information on results incl. tables'.
Any other information on results incl. tables
Table 1: Number of revertants per plate (mean of 3 plates), first experiment [1b]
|
|
TA 98 |
TA100 |
TA 102 |
TA 1535 |
TA1537 |
||||||||||||||
Conc. |
— MA |
Cytotoxic Precipitates (yes/no) |
+ MA |
Cytotoxic Precipitates (yes/no) |
— MA |
Cytotoxic Precipitates (yes/no) |
+ MA |
Cytotoxic Precipitates (yes/no) |
— MA |
Cytotoxic Precipitates (yes/no) |
+ MA |
Cytotoxic Precipitates (yes/no) |
— MA |
Cytotoxic Precipitates (yes/no) |
+ MA |
Cytotoxic Precipitates (yes/no) |
— MA |
Cytotoxic Precipitates (yes/no) |
+ MA |
Cytotoxic Precipitates (yes/no) |
demin water |
15 |
no |
15 |
no |
71 |
no |
73 |
no |
264 |
no |
275 |
no |
10 |
no |
18 |
no |
7 |
no |
9 |
no |
0* |
13 |
no |
16 |
no |
60 |
no |
69 |
no |
259 |
no |
277 |
no |
17 |
no |
13 |
no |
9 |
no |
8 |
no |
50 |
11 |
no |
13 |
no |
63 |
no |
61 |
no |
280 |
no |
275 |
no |
12 |
no |
13 |
no |
8 |
no |
6 |
no |
150 |
14 |
no |
11 |
no |
65 |
no |
75 |
no |
309 |
no |
309 |
no |
12 |
no |
11 |
no |
4 |
no |
7 |
no |
500 |
10 |
no |
9 |
no |
67 |
no |
61 |
no |
317 |
no |
312 |
no |
11 |
no |
11 |
no |
8 |
no |
8 |
no |
1500 |
11 |
no |
9 |
no |
91 |
no |
87 |
no |
293 |
no |
285 |
no |
9 |
no |
8 |
no |
7 |
no |
6 |
no |
5000 |
7 |
no |
11 |
no |
110 |
no |
121 |
no |
291 |
no |
293 |
no |
10 |
no |
11 |
no |
6 |
no |
7 |
no |
Positive control |
s.g. |
no |
98 |
no |
s.g. |
no |
s.g. |
no |
605 |
no |
s.g. |
no |
221 |
no |
175 |
no |
232 |
no |
168 |
no |
*solvent/vehicle control with DMSO
Table 1b: Number of revertants per plate (mean of 3 plates), first experiment [1d]
|
|
TA 1537 |
||
Conc. |
— MA |
Cytotoxic Precipitates (yes/no) |
+ MA |
Cytotoxic Precipitates (yes/no) |
demin water |
5 |
no |
5 |
no |
0* |
4 |
no |
6 |
no |
50 |
5 |
no |
5 |
no |
150 |
5 |
no |
5 |
no |
500 |
4 |
no |
4 |
no |
1500 |
4 |
no |
4 |
no |
5000 |
2 |
no |
2 |
no |
Positive control |
91 |
no |
149 |
no |
Table 2: Number of revertants per plate (mean of 3 plates), second experiment
|
|
TA 98 |
TA100 |
TA 102 |
TA 1535 |
TA1537 |
||||||||||||||
Conc. |
— MA |
Cytotoxic Precipitates (yes/no) |
+ MA |
Cytotoxic Precipitates (yes/no) |
— MA |
Cytotoxic Precipitates (yes/no) |
+ MA |
Cytotoxic Precipitates (yes/no) |
— MA |
Cytotoxic Precipitates (yes/no) |
+ MA |
Cytotoxic Precipitates (yes/no) |
— MA |
Cytotoxic Precipitates (yes/no) |
+ MA |
Cytotoxic Precipitates (yes/no) |
— MA |
Cytotoxic Precipitates (yes/no) |
+ MA |
Cytotoxic Precipitates (yes/no) |
demin water |
20 |
no |
19 |
no |
91 |
no |
89 |
no |
371 |
no |
376 |
no |
10 |
no |
10 |
no |
5 |
no |
8 |
no |
0* |
18 |
no |
20 |
no |
81 |
no |
83 |
no |
355 |
no |
365 |
no |
9 |
no |
12 |
no |
6 |
no |
8 |
no |
78 |
15 |
no |
16 |
no |
61 |
no |
61 |
no |
371 |
no |
357 |
no |
7 |
no |
6 |
no |
6 |
no |
6 |
no |
156 |
15 |
no |
16 |
no |
69 |
no |
65 |
no |
365 |
no |
371 |
no |
8 |
no |
7 |
no |
7 |
no |
6 |
no |
313 |
17 |
no |
16 |
no |
80 |
no |
77 |
no |
363 |
no |
365 |
no |
7 |
no |
7 |
no |
5 |
no |
4 |
no |
1250 |
16 |
no |
16 |
no |
85 |
no |
83 |
no |
357 |
no |
352 |
no |
6 |
no |
5 |
no |
3 |
no |
3 |
no |
2500 |
15 |
no |
16 |
no |
57 |
no |
63 |
no |
304 |
no |
320 |
no |
3 |
no |
5 |
no |
0 |
no |
1 |
no |
5000 |
0 |
no |
0 |
no |
4 |
no |
2 |
no |
269 |
no |
275 |
no |
1 |
no |
1 |
no |
0 |
no |
0 |
no |
Positive control |
s.g. |
no |
111 |
no |
s.g. |
no |
s.g. |
no |
765 |
no |
872 |
no |
248 |
no |
167 |
no |
152 |
no |
160 |
no |
*solvent/vehicle control with water
Concurrent controls
DMSO Experiment 1b/Vehicle control
Strain |
TA98 |
TA100 |
TA102 |
TA1535 |
TA1537 |
|||||
Induction |
-S9 |
+S9 |
-S9 |
+S9 |
-S9 |
+S9 |
-S9 |
+S9 |
-S9 |
+S9 |
Repl.1 |
11 |
15 |
60 |
62 |
256 |
288 |
15 |
15 |
11 |
8 |
Repl.2 |
14 |
13 |
62 |
80 |
272 |
264 |
17 |
10 |
8 |
9 |
Repl.3 |
14 |
19 |
58 |
66 |
248 |
280 |
18 |
14 |
8 |
8 |
Mean |
13 |
16 |
60 |
69 |
259 |
277 |
17 |
13 |
9 |
8 |
sd |
1.7 |
3.1 |
2.0 |
9.5 |
12.2 |
12.2 |
1.5 |
2.6 |
1.7 |
0.6 |
Demin Water Experiment 1b/Vehicle control
Strain |
TA98 |
TA100 |
TA102 |
TA1535 |
TA1537 |
|||||
Induction |
-S9 |
+S9 |
-S9 |
+S9 |
-S9 |
+S9 |
-S9 |
+S9 |
-S9 |
+S9 |
Repl.1 |
15 |
18 |
62 |
70 |
280 |
9 |
17 |
8 |
9 |
9 |
Repl.2 |
13 |
10 |
78 |
72 |
248 |
272 |
9 |
17 |
8 |
10 |
Repl.3 |
16 |
17 |
72 |
76 |
264 |
272 |
12 |
19 |
6 |
8 |
Mean |
15 |
15 |
71 |
73 |
264 |
275 |
10 |
18 |
7 |
9 |
sd |
1.5 |
4.4 |
8.1 |
3.1 |
16.0 |
4.6 |
1.7 |
1.2 |
1.2 |
1.0 |
Positive control Experiment 1b
Strain |
TA98 |
TA100 |
TA102 |
TA1535 |
TA1537 |
|||||
Induction |
-S9 |
+S9 |
-S9 |
+S9 |
-S9 |
+S9 |
-S9 |
+S9 |
-S9 |
+S9 |
Substance |
NPD |
BaP |
Na-azide |
2-AA |
NPD |
2-AA |
Na-azide |
2-AA |
NPD |
2-AA |
Repl.1 |
s.g. |
100 |
s.g. |
s.g. |
576 |
s.g. |
220 |
180 |
232 |
160 |
Repl.2 |
s.g. |
80 |
s.g. |
s.g. |
608 |
s.g. |
228 |
172 |
240 |
192 |
Repl.3 |
s.g. |
114 |
s.g. |
s.g. |
632 |
s.g. |
216 |
172 |
224 |
152 |
Mean |
-- |
98 |
-- |
-- |
605 |
-- |
221 |
175 |
232 |
168 |
sd |
-- |
17.1 |
-- |
-- |
28.1 |
-- |
6.1 |
4.6 |
8.0 |
21.2 |
f(l) |
>2 |
6.13 |
>2 |
>2 |
2.34 |
>2 |
22.10 |
13.46 |
25.78 |
21.00 |
Rev. abs. |
-- |
82 |
-- |
-- |
346 |
-- |
211 |
162 |
223 |
160 |
DMSO Experiment 1d/Vehicle control
Strain |
TA1537 |
|
Induction |
-S9 |
+S9 |
Repl. 1 |
4 |
5 |
Repl. 2 |
3 |
7 |
Repl. 3 |
5 |
6 |
Mean |
5 |
6 |
SD |
1.0 |
1.0 |
Demin Water Experiment 1d/Vehicle control
Strain |
TA1537 |
|
Induction |
-S9 |
+S9 |
Repl. 1 |
4 |
6 |
Repl. 2 |
5 |
4 |
Repl. 3 |
6 |
5 |
Mean |
5 |
5 |
SD |
1.0 |
1.0 |
Positive control Experiment 1d
Strain |
TA1537 |
|
Induction |
-S9 |
+S9 |
Substance |
Na-Azide |
2-AA |
Repl. 1 |
96 |
160 |
Repl. 2 |
92 |
148 |
Repl. 3 |
84 |
140 |
Mean |
91 |
149 |
SD |
6.1 |
10.1 |
f(l) |
22.75 |
24.83 |
Rev. abs. |
87 |
143 |
s.g.= strong growth, too strong for counting of revertants
f(l) = increase factor
Rev.abs. = absolute revertants
Demin. Water Experiment 2/Vehicle control
Strain |
TA98 |
TA100 |
TA102 |
TA1535 |
TA1537 |
|||||
Induction |
-S9 |
+S9 |
-S9 |
+S9 |
-S9 |
+S9 |
-S9 |
+S9 |
-S9 |
+S9 |
Repl.1 |
19 |
20 |
96 |
96 |
352 |
368 |
9 |
10 |
6 |
9 |
Repl.2 |
20 |
19 |
92 |
92 |
384 |
384 |
12 |
10 |
5 |
7 |
Repl.3 |
20 |
19 |
84 |
80 |
376 |
376 |
10 |
11 |
4 |
7 |
Mean |
20 |
19 |
91 |
89 |
371 |
376 |
10 |
10 |
5 |
8 |
sd |
0.6 |
0.6 |
6.1 |
8.3 |
16.7 |
8.0 |
1.5 |
0.6 |
1.0 |
1.2 |
DMSO Experiment 2/Vehicle control
Strain |
TA98 |
TA100 |
TA102 |
TA1535 |
TA1537 |
|||||
Induction |
-S9 |
+S9 |
-S9 |
+S9 |
-S9 |
+S9 |
-S9 |
+S9 |
-S9 |
+S9 |
Repl.1 |
18 |
19 |
76 |
80 |
352 |
360 |
9 |
11 |
6 |
9 |
Repl.2 |
18 |
20 |
80 |
84 |
360 |
368 |
8 |
15 |
6 |
6 |
Repl.3 |
18 |
21 |
88 |
84 |
352 |
368 |
10 |
10 |
6 |
8 |
Mean |
18 |
20 |
81 |
83 |
355 |
365 |
9 |
12 |
6 |
8 |
sd |
0.0 |
1.0 |
6.1 |
2.3 |
4.6 |
4.6 |
1.0 |
2.6 |
0.0 |
1.5 |
Positive control Experiment 2
Strain |
TA98 |
TA100 |
TA102 |
TA1535 |
TA1537 |
|||||
Induction |
-S9 |
+S9 |
-S9 |
+S9 |
-S9 |
+S9 |
-S9 |
+S9 |
-S9 |
+S9 |
Substance |
NPD |
BaP |
Na-azide |
2-AA |
NPD |
2-AA |
Na-azide |
2-AA |
NPD |
2-AA |
Repl.1 |
s.g. |
112 |
s.g. |
s.g. |
752 |
856 |
240 |
164 |
156 |
168 |
Repl.2 |
s.g. |
112 |
s.g. |
s.g. |
768 |
864 |
264 |
168 |
132 |
168 |
Repl.3 |
s.g. |
108 |
s.g. |
s.g. |
776 |
896 |
240 |
168 |
168 |
144 |
Mean |
-- |
111 |
-- |
-- |
765 |
872 |
248 |
167 |
152 |
160 |
sd |
-- |
2.3 |
-- |
-- |
12.2 |
21.2 |
13.9 |
2.3 |
18.3 |
13.9 |
f(l) |
>2 |
5.55 |
>2 |
>2 |
2.15 |
2.39 |
24.80 |
13.92 |
25.33 |
20.00 |
Rev. abs. |
-- |
91 |
-- |
-- |
410 |
507 |
238 |
155 |
146 |
152 |
s.g.= strong growth, too strong for counting of revertants
f(l) = increase factor
Rev.abs. = absolute revertants
Applicant's summary and conclusion
- Conclusions:
- There was no evidence of induced mutant colonies over background, when bis(glycinato)copper was tested with and without metabolic activation up to the recommended limit concentration (5000 µg/plate).
- Executive summary:
In a reverse gene mutation assay in bacteria according to OECD guideline 471 and EU Method B.14 (Version Commission Directive 92/69/EEC), strains TA1535, TA 1537, TA 102, TA 100 and TA 98 of S. typhimurium were exposed to bis(glycinato)copper. The test was performed with concentrations up to the recommended limit concentration of 5000 µg/plate in the absence and the presence of mammalian metabolic activation (S9 -mix).
No evidence of biologically significant mutagenic activity of the test item was found in the presence and absence of metabolic activation, up to the limit concentration of 5000 µg/plate. The positive controls induced the appropriate responses in the corresponding strains and activity of metabolizing system was confirmed.
There was no evidence of induced mutant colonies over background. Based on the results of this study, it is concluded that bis(glycinato)copper is not mutagenic in the Salmonella typhimurium strains TA98, TA100, TA102, TA1535 and TA1537 in the presence and absence of metabolic activation under the experimental conditions in this study.
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