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EC number: 407-870-4 | CAS number: 97384-48-0 CITROWANIL B
- 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
- Remarks:
- Type of genotoxicity: gene mutation
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Study period:
- 2010-09-28 to 2011-01-10
- Reliability:
- 2 (reliable with restrictions)
- Rationale for reliability incl. deficiencies:
- other: see 'Remark'
- Remarks:
- Test according to EU-Method B.13/14 adopted 31. May, 2008 "Mutagenicity - Reverse mutation test using bacteria", Test according to OECD Guideline for Testing of Chemicals Part 471, adopted 21. Jul. 1997 "Bacterial Reverse Mutation Test", Test according to "Revised OECD Principles of Good Laboratory Practice" (Paris, 1997).
Data source
Reference
- Reference Type:
- study report
- Title:
- Unnamed
- Year:
- 2 011
- Report date:
- 2011
Materials and methods
Test guidelineopen allclose all
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 471 (Bacterial Reverse Mutation Assay)
- Deviations:
- no
- Qualifier:
- according to guideline
- Guideline:
- EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)
- Deviations:
- no
- GLP compliance:
- yes (incl. QA statement)
- Type of assay:
- bacterial reverse mutation assay
Test material
- Reference substance name:
- CITROWANIL B, colourless to pale yellow liquid
- IUPAC Name:
- CITROWANIL B, colourless to pale yellow liquid
- Reference substance name:
- 2-benzyl-2-methyl-3-butennitril
- IUPAC Name:
- 2-benzyl-2-methyl-3-butennitril
- Reference substance name:
- 2-benzyl-2-methyl-3-butenitrile
- EC Number:
- 407-870-4
- EC Name:
- 2-benzyl-2-methyl-3-butenitrile
- Cas Number:
- 97384-48-0
- Molecular formula:
- C12H13N
- IUPAC Name:
- 2-benzyl-2-methylbut-3-enenitrile
- Details on test material:
- Citrowanil B; C12H13N
Constituent 1
Constituent 2
Constituent 3
Method
- Target gene:
- TA97a: hisD6610, uvrB, pKM 101, rfa
TA98: hisD3052, uvrB, pKM 101, rfa
TA100: hisG46, uvrB, pKM 101, rfa
TA102: hisG428,pKM 101, rfa
TA1535: hisG46, uvrB, rfa
Species / strainopen allclose all
- Species / strain / cell type:
- other: S. typhimurium TA97a
- Additional strain / cell type characteristics:
- other: hisD6610, uvrB, pKM 101, rfa
- Species / strain / cell type:
- S. typhimurium TA 102
- Additional strain / cell type characteristics:
- other: hisG428,pKM 101, rfa
- Species / strain / cell type:
- S. typhimurium TA 100
- Additional strain / cell type characteristics:
- other: hisG46, uvrB, pKM 101, rfa
- Species / strain / cell type:
- S. typhimurium TA 98
- Additional strain / cell type characteristics:
- other: hisD3052, uvrB, pKM 101, rfa
- Species / strain / cell type:
- S. typhimurium TA 1535
- Additional strain / cell type characteristics:
- other: hisG46, uvrB, rfa
- Metabolic activation:
- with and without
- Metabolic activation system:
- S9-Mix with 4% S9, produced from the livers of male Spraque-Dawley rats wich were treated with 500 mg Aroclor 1254/kg body weight intraperitoneally
- Test concentrations with justification for top dose:
- First Experiment: 4998 / 1499 / 500 / 150 / 50 µg/plate
Second Experiment: 501 / 150 / 50 / 15 / 5 µg/plate
Third Experiment: 504 / 252 / 126 / 63 / 32 / 16 / 8 µg/plate - Vehicle / solvent:
- DMSO
Controlsopen allclose all
- Untreated negative controls:
- yes
- Remarks:
- water
- Negative solvent / vehicle controls:
- yes
- Remarks:
- DMSO
- Positive controls:
- yes
- Positive control substance:
- benzo(a)pyrene
- Remarks:
- Migrated to IUCLID6: 20 µg/plate in DMSO for TA98 with metabolic activation
- Positive controls:
- yes
- Positive control substance:
- sodium azide
- Remarks:
- Migrated to IUCLID6: 1 µg/plate in water for TA100, TA1535 without metabolic activation
- Positive controls:
- yes
- Positive control substance:
- other: 1 µg/plate 2-Amino-anthracene in DMSO for TA97a, TA100, TA102, TA1535 with metabolic activation
- Positive controls:
- yes
- Positive control substance:
- other: 20 µg/plate 4-Nitro-1,2-phenylene diaminein DMSO for TA97a, TA98, TA102 without metabolic activation
- Details on test system and experimental conditions:
- Per strain and dose, four plates with and four plates without S9 were used.
The first and the second experiment were carried out according to the plate incorporation method, the third experiment was carried out according to the Pre-incubation method.
Each test item solution was membrane filtrated before use to accomplish sterility.
Plate incorporation method:
0.1 mL of the appropriate solution of the test item was given into a sterile tube. After mixing with 0.1 mL overnight culture of the respective strain and 0.5 mL phosphate buffer (treatement without S9) or 0.5 mL S9 mix (treatement with S9), 2 mL Top-Agar, containing 10 mL 0.5 mMol histidine-biotin-solution per 100 mL, were added.
Pre-incubation method:
0.1 mL of the appropriate solution of the test item was given into a sterile tube. After mixing with 0.1 mL overnight culture of the respective strain and 0.5 mL phosphate buffer (treatement without S9) or 0.5 mL S9 mix (treatement with S9) were added. The mixture was incubated in an incubating chamber at 37°C for 20 minutes. During this time the vessles were aerated through careful shaking. Then 2 mL Top-Agar, containing 10 mL 0.5 mMol histidine-biotin-solution per 100 mL, was added.
In each case the mixture was vortexed gently, then poured on a minimal glucose plate and distributed evenly, rotating the plate. The plates were closed and left to harden for a few minutes, then inverted and placed in the dark incubator at 37°C. - Evaluation criteria:
- The colonies were counted visually after 48 hours incubation, the numbers were recorded.
A test item is considered to have mutagenic potential, if a significant, reproducible increase of revertant colonies per plate (increase factor f(I) ≥ 2) in at least one strain can be observed. A concentration-releated increase over the range tested can also be taken as a sign of mutagenic activity. - Statistics:
- A spreadsheet software (Microsoft Excel®) was used to calculate mean values and standard deviations of each treatment, solvent control and positive control.
The increase factor f(I) of revertant induction (mean revetants divided by mean spontaneous revertants [vehicle control]) and the absolute number of revertants (mean revetants less mean spontaneous revertants [vehicle control]) were calculated, too.
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:
- other: yes at 4998 / 1499 µg/plate, no at 500 / 150 / 50 µg/plate
- Vehicle controls validity:
- valid
- Untreated negative 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:
- other: yes at 4998 / 1499 µg/plate, no at 500 / 150 / 50 µg/plate
- Vehicle controls validity:
- valid
- Untreated negative 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:
- other: yes at 4998 / 1499 µg/plate, no at 500 / 150 / 50 µg/plate
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- valid
- Positive controls validity:
- valid
- Species / strain:
- S. typhimurium TA 102
- Metabolic activation:
- with and without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- other: yes at 4998 / 1499 µg/plate, no at 500 / 150 / 50 µg/plate
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- valid
- Positive controls validity:
- valid
- Species / strain:
- other: S. typhimurium TA97a
- Metabolic activation:
- with and without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- no cytotoxicity
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- valid
- Positive controls validity:
- valid
- Remarks on result:
- other: other: Plate incorporation and Pre-incubation method
- Remarks:
- Migrated from field 'Test system'.
Any other information on results incl. tables
Survey of the Findings:
The mean revertant values of the four replicates are presented in the following table.
Exp. |
Strain |
TA97a |
TA98 |
TA100 |
TA102 |
TA1535 |
||||||
Induction |
-S9 |
+S9 |
-S9 |
+S9 |
-S9 |
+S9 |
-S9 |
+S9 |
-S9 |
+S9 |
||
1 |
H2O |
Mean |
124 |
115 |
11 |
17 |
128 |
113 |
218 |
233 |
14 |
12 |
DMSO |
Mean |
112 |
105 |
11 |
14 |
112 |
117 |
191 |
294 |
12 |
13 |
|
Pos. Contr. |
Mean f(I) |
768 6.9 |
789 7.5 |
752 68.4 |
142 10.1 |
633 5.0 |
496 4.2 |
817 4.3 |
657 2.2 |
242 17.3 |
144 11.1 |
|
4998 µg/pl. |
Mean f(I) |
110 0.98 |
67 0.64 |
1 0.09 |
0 0.00 |
0 0.00 |
0 0.00 |
20 0.10 |
0 0.00 |
0 0.00 |
0 0.00 |
|
1499 µg/pl. |
Mean f(I) |
57 0.51 |
115 1.10 |
0 0.00 |
0 0.00 |
0 0.00 |
27 0.23 |
23 0.12 |
15 0.05 |
0 0.00 |
0 0.00 |
|
500 µg/pl. |
Mean f(I) |
101 0.90 |
111 1.06 |
9 0.82 |
10 0.71 |
123 1.10 |
129 1.10 |
229 1.20 |
217 0.74 |
10 0.83 |
15 1.15 |
|
150 µg/pl. |
Mean f(I) |
113 1.01 |
132 1.26 |
10 0.91 |
11 0.79 |
130 1.16 |
130 1.11 |
206 1.08 |
241 0.82 |
11 0.92 |
12 0.92 |
|
50 µg/pl. |
Mean f(I) |
116 1.04 |
113 1.08 |
9 0.82 |
12 0.86 |
125 1.12 |
111 0.95 |
282 1.48 |
234 0.80 |
13 1.08 |
13 1.00 |
|
|
||||||||||||
2 |
H2O |
Mean |
112 |
101 |
11 |
15 |
99 |
115 |
208 |
200 |
14 |
14 |
DMSO |
Mean |
105 |
97 |
11 |
16 |
90 |
111 |
253 |
176 |
10 |
12 |
|
Pos. Contr. |
Mean f(I) |
310 3.0 |
414 4.3 |
305 27.7 |
260 16.3 |
674 6.8 |
1084 9.8 |
639 2.5 |
796 4.5 |
226 16.1 |
122 10.2 |
|
501 µg/pl. |
Mean f(I) |
114 1.09 |
107 1.10 |
10 0.91 |
12 0.75 |
108 1.20 |
99 0.89 |
151 0.60 |
139 0.79 |
10 1.00 |
13 1.08 |
|
150 µg/pl. |
Mean f(I) |
117 1.11 |
125 1.29 |
11 1.00 |
15 0.94 |
100 1.11 |
148 1.33 |
190 0.75 |
176 1.00 |
13 1.30 |
9 0.75 |
|
50 µg/pl. |
Mean f(I) |
109 1.04 |
99 1.02 |
11 1.00 |
12 0.75 |
126 1.40 |
83 0.75 |
158 0.62 |
168 0.95 |
13 1.30 |
14 1.17 |
|
15 µg/pl. |
Mean f(I) |
110 1.05 |
91 0.94 |
11 1.00 |
13 0.81 |
88 0.98 |
113 1.02 |
179 0.71 |
182 1.03 |
11 1.10 |
18 1.50 |
|
5 µg/pl. |
Mean f(I) |
94 0.90 |
105 1.08 |
12 1.09 |
12 0.75 |
111 1.23 |
84 0.76 |
213 0.84 |
205 1.16 |
13 1.30 |
16 1.33 |
|
|
||||||||||||
3 |
H2O |
Mean |
110 |
108 |
11 |
16 |
78 |
127 |
199 |
174 |
16 |
14 |
DMSO |
Mean |
105 |
105 |
14 |
11 |
98 |
106 |
196 |
186 |
17 |
17 |
|
Pos. Contr. |
Mean f(I) |
320 3.1 |
303 2.9 |
231 16.5 |
117 10.6 |
254 3.3 |
221 2.1 |
886 4.5 |
514 2.8 |
237 14.8 |
208 12.2 |
|
504 µg/pl. |
Mean f(I) |
115 1.10 |
135 1.29 |
6 0.43 |
9 0.82 |
29 0.30 |
91 0.86 |
222 1.13 |
224 1.2 |
9 0.53 |
10 0.59 |
|
252 µg/pl. |
Mean f(I) |
106 1.01 |
104 0.99 |
5 0.36 |
12 1.09 |
29 0.30 |
84 0.79 |
230 1.17 |
188 1.01 |
13 0.76 |
14 0.82 |
|
126 µg/pl. |
Mean f(I) |
121 1.15 |
111 1.06 |
8 0.57 |
13 1.18 |
79 0.81 |
104 0.98 |
268 1.37 |
192 1.03 |
10 0.59 |
19 1.12 |
|
63 µg/pl. |
Mean f(I) |
111 1.06 |
112 1.07 |
10 0.71 |
14 1.27 |
99 1.01 |
126 1.19 |
159 0.81 |
174 0.94 |
11 0.65 |
16 0.94 |
|
32 µg/pl. |
Mean f(I) |
108 1.03 |
107 1.02 |
10 0.71 |
13 1.18 |
108 1.10 |
88 0.83 |
211 1.08 |
258 1.39 |
11 0.65 |
14 0.82 |
|
18 µg/pl |
Mean f(I) |
112 1.07 |
108 1.03 |
9 0.64 |
18 1.64 |
88 0.90 |
82 0.77 |
219 1.12 |
228 1.23 |
12 0.71 |
14 0.82 |
|
8 µg/pl |
Mean f(I) |
112 1.07 |
113 1.08 |
8 0.57 |
16 1.45 |
107 1.09 |
110 1.04 |
218 1.11 |
156 0.84 |
11 0.65 |
12 0.71 |
f(I) = increase factor
Applicant's summary and conclusion
- Conclusions:
- Interpretation of results (migrated information):
negative without metabolic activation
negative with metabolic activation
CITROWANIL B is not mutagenic in the Bacterial Reverse Mutation Test using Salmonella typhimurium, strains TA97a, TA98, TA100, TA102 and TA1535. - Executive summary:
Three valid experiments were performed to evaluate the mutagenic potential of CITROWANIL B in the Bacterial Reverse Mutation Test.
First Experiment:
Five concentrations of the test item, dissolved in dimethyl sulfoxid (DMSO) (ranging from 4998 to 50 µg/plate) were used. Five genetically manipulated strains ofSalmonella typhimurium(TA97a, TA98, TA100, TA102 and TA1535) were exposed to the test item both in the presence and in the absence of a metabolic activation system (S9) for 48 hours using the plate incorporation method.
Signs of toxicity towards the bacteria could be observed in the two highest concentrations (4998 and 1499 µg/plate).
None of the three last concentrations (500, 150 and 50 µg/plate) caused a significant increase in the numbers of revertant colonies in the tested strains. The test item didn’t show any mutagenic effect in the first experiment.
The sterility control and the determination of the titre didn’t show any inconsistencies. The determined values for the spontaneous revertants of the negative controls were within the normal range. All positive controls showed mutagenic effects with and without metabolic activation.
Second Experiment:
Because of the results of the first experiment, a second experiment was performed, using five concentrations of the test item (ranging from 501 to 5 µg/plate), using the plate incorporation method, too.
The test item didn’t show mutagenic effects in the second experiment, either.
No signs of toxicity towards the bacteria could be observed.
The sterility control and the determination of the titre didn’t show any inconsistencies. The determined values for the spontaneous revertants of the negative controls were within the normal range. All positive controls showed mutagenic effects with and without metabolic activation.
Third Experiment:
To verify the first and the second experiment, a third experiment was performed, using seven concentrations of the test item (ranging from 504 to 8 µg/plate) and a modification in study performance(pre-incubation method).
The test item didn’t show mutagenic effects in the third experiment, either.
No signs of toxicity towards the bacteria could be observed.
The sterility control and the determination of the titre didn’t show any inconsistencies. The determined values for the spontaneous revertants of the negative controls were within the normal range. All positive controls showed mutagenic effects with and without metabolic activation.
Under the conditions of the test, the test item didn’t show mutagenic effects in Salmonella typhimurium, strains TA97a, TA98, TA100, TA102 and TA1535.
Therefore, no concentration-effect relationship could be determined.
The test item CITROWANIL B is considered as “not mutagenic under the conditions of the test”.
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