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EC number: 947-664-6 | CAS number: -
- 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:
- 09 to 27 Februray 2018
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
- Remarks:
- GLP study conducted according to OECD Guideline 471 without any deviation
Data source
Reference
- Reference Type:
- study report
- Title:
- Unnamed
- Year:
- 2 018
- Report date:
- 2018
Materials and methods
Test guidelineopen allclose all
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 471 (Bacterial Reverse Mutation Assay)
- Version / remarks:
- dated 21 July 1997
- Deviations:
- no
- Qualifier:
- according to guideline
- Guideline:
- EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)
- Version / remarks:
- dated 30 May 2008
- Deviations:
- no
- Qualifier:
- according to guideline
- Guideline:
- EPA OPPTS 870.5100 - Bacterial Reverse Mutation Test (August 1998)
- Version / remarks:
- August 1998
- Deviations:
- no
- Qualifier:
- according to guideline
- Guideline:
- other: ENV/EPOC(97)4 OECD Guideline for the Testing of Chemicals, Proposal for Replacement of Guidelines 471 and 472 Bacterial Reverse Mutation Test
- Deviations:
- no
- Qualifier:
- according to guideline
- Guideline:
- other: EEC Directive 2000/32, L1362000, Annex 4D, dated May 19, 2000
- Deviations:
- no
- Qualifier:
- according to guideline
- Guideline:
- other: The Japanese Ministry of Economy Trade and Industry (METI), Ministry of Health, Labour and Welfare (MHLW) and Ministry of the Environment (MOE) Guidelines of 31 March 2011
- Deviations:
- no
- Principles of method if other than guideline:
- Not applicable
- GLP compliance:
- yes (incl. QA statement)
- Remarks:
- 14 September 2015
- Type of assay:
- bacterial reverse mutation assay
Test material
- Reference substance name:
- 7,11-dimethyl-3-methylenedodeca-1,6,10-triene
- EC Number:
- 278-628-1
- EC Name:
- 7,11-dimethyl-3-methylenedodeca-1,6,10-triene
- Cas Number:
- 77129-48-7
- Molecular formula:
- C15H24
- IUPAC Name:
- 7,11-dimethyl-3-methylidenedodeca-1,6,10-triene
- Reference substance name:
- (R*,R*)-α,4-dimethyl-α-(4-methyl-3-pentenyl)cyclohex-3-ene-1-methanol
- EC Number:
- 208-205-9
- EC Name:
- (R*,R*)-α,4-dimethyl-α-(4-methyl-3-pentenyl)cyclohex-3-ene-1-methanol
- Cas Number:
- 515-69-5
- Molecular formula:
- C15H26O
- IUPAC Name:
- 6-methyl-2-(4-methylcyclohex-3-en-1-yl)hept-5-en-2-ol
- Reference substance name:
- [3S-[3α,6α(R*)]-tetrahydro-2,2,6-trimethyl-6-(4-methyl-3-cyclohexen-1-yl)-2H-pyran-3-ol
- EC Number:
- 245-086-2
- EC Name:
- [3S-[3α,6α(R*)]-tetrahydro-2,2,6-trimethyl-6-(4-methyl-3-cyclohexen-1-yl)-2H-pyran-3-ol
- Cas Number:
- 22567-36-8
- Molecular formula:
- C15H26O2
- IUPAC Name:
- 2,2,6-trimethyl-6-(4-methylcyclohex-3-en-1-yl)tetrahydro-2H-pyran-3-ol
- Reference substance name:
- 7-ethyl-1,4-dimethylazulene
- EC Number:
- 208-449-6
- EC Name:
- 7-ethyl-1,4-dimethylazulene
- Cas Number:
- 529-05-5
- Molecular formula:
- C14H16
- IUPAC Name:
- 7-ethyl-1,4-dimethylazulene
- Reference substance name:
- [2S-[2α,5β(R*)]]-tetrahydro-α,α,5-trimethyl-5-(4-methyl-3-cyclohexen-1-yl)furan-2-methanol
- EC Number:
- 247-509-6
- EC Name:
- [2S-[2α,5β(R*)]]-tetrahydro-α,α,5-trimethyl-5-(4-methyl-3-cyclohexen-1-yl)furan-2-methanol
- Cas Number:
- 26184-88-3
- Molecular formula:
- C15H26O2
- IUPAC Name:
- 2-[5-methyl-5-(4-methylcyclohex-3-en-1-yl)tetrahydrofuran-2-yl]propan-2-ol
- Reference substance name:
- [S-(R*,R*)]-dihydro-2,2,6-trimethyl-6-(4-methyl-3-cyclohexen-1-yl)-2H-pyran-3(4H)-one
- EC Number:
- 245-087-8
- EC Name:
- [S-(R*,R*)]-dihydro-2,2,6-trimethyl-6-(4-methyl-3-cyclohexen-1-yl)-2H-pyran-3(4H)-one
- Cas Number:
- 22567-38-0
- Molecular formula:
- C15H24O2
- IUPAC Name:
- 2,2,6-trimethyl-6-(4-methylcyclohex-3-en-1-yl)dihydro-2H-pyran-3(4H)-one
- Reference substance name:
- 2,6,10-trimethyldodeca-2,6,9,11-tetraene
- EC Number:
- 207-948-6
- EC Name:
- 2,6,10-trimethyldodeca-2,6,9,11-tetraene
- Cas Number:
- 502-61-4
- Molecular formula:
- C15H24
- IUPAC Name:
- (3E,6E)-3,7,11-trimethyldodeca-1,3,6,10-tetraene
- Reference substance name:
- (Z)-en-yn-dicycloether
- Cas Number:
- 4575-53-5
- Molecular formula:
- C13H14O2
- IUPAC Name:
- (Z)-en-yn-dicycloether
- Test material form:
- liquid
- Details on test material:
- Test item: Blue chamomile oil
Batch No.: 2102919-85304
EC No.: 926-224-7
CAS No.: 8002-66-2
Content: 100 % UVCB
Physical state: Dark blue liquid
Storage conditions: Original aluminium bottle, ambient temperature (10 °C to 30 °C)
Expiry date: October 02, 2019
Certificate: Specifications, Silvestris, 2144 Kerepes, Hungary
Constituent 1
Constituent 2
Constituent 3
Constituent 4
Constituent 5
Constituent 6
Constituent 7
Constituent 8
- Specific details on test material used for the study:
- Storage Conditions:Room temperature
Method
- Target gene:
- he histidine dependent strains are derived from Salmonella typhimurium strain LT2 through mutations in the histidine locus. Additionally due to the "deep rough" (rfa-minus) mutation they possess a faulty lipopolysaccharide envelope which enables substances to penetrate the cell wall more easily. A further mutation causes a reduction in the activity of an excision repair system. The latter alteration includes mutational processes in the nitrate reductase and biotin genes produced in a UV-sensitive area of the gene named "uvrB-minus". In the strains TA 98 and TA 100 the R-factor plasmid pKM 101 carries the ampicillin resistance marker.
Strain Escherichia coli WP2 and its derivatives carry the same defect in one of the genes for tryptophan biosynthesis. Tryptophan-independent (Trp+) mutants (revertants) can arise either by a base change at the site of the original alteration or by a base change elsewhere in the chromosome so that the original defect is suppressed. This second possibility can occur in several different ways so that the system seems capable of detecting all types of mutagen which substitute one base for another. Additionally, the uvrA derivative is deficient in the DNA repair process (excision repair damage). Such a repair-deficient strain may be more readily mutated by agents.
Species / strain
- Species / strain / cell type:
- S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and E. coli WP2
- Details on mammalian cell type (if applicable):
- Not applicable
- Additional strain / cell type characteristics:
- not applicable
- Cytokinesis block (if used):
- Not applicable
- Metabolic activation:
- with and without
- Metabolic activation system:
- rat liver homogenate metabolizing system (10% liver S9 in standard co-factors)
- Test concentrations with justification for top dose:
- According to the standard limit concentrations recommended in the regulatory guidelines 5000 µg/plate was applied as the highest concentration in the pre-experiment.
Pre-Experiment/Experiment I: 3; 10; 33; 100; 333; 1000; 2500; and 5000 µg/plate
Experiment II:
Strains TA 100: 0.3; 1; 3; 10; 33; 100; 333; and 1000 µg/plate
The remaining strains: 33; 100; 333; 1000; 2500; and 5000 µg/plate - Vehicle / solvent:
- On the day of the experiment, the test item was dissolved in DMSO (purity > 99 %). The solvent has been chosen according to its solubility properties and its relative nontoxicity to the bacteria (Maron et al.; 1981)
Controls
- Untreated negative controls:
- yes
- Negative solvent / vehicle controls:
- yes
- Remarks:
- DMSO
- Positive controls:
- yes
- Positive control substance:
- sodium azide
- methylmethanesulfonate
- other: 4-nitro-o-phenylene-diamine and 2-aminoanthracene
- Details on test system and experimental conditions:
- SOURCE OF TEST SYSTEM: The bacterial strains TA 1535, TA 1537, TA 98, TA 100, and WP2 uvrA were obtained from Trinova Biochem GmbH (35394 Gießen, Germany).
METHOD OF APPLICATION:
EXPERIMENT: Experiment I (Plate Incorporation)
The following materials were mixed in a test tube and poured onto the selective agar plates:
100 µL Test solution at each dose level (solvent or reference mutagen solution (positive control)),
500 µL S9 mix (for test with metabolic activation) or S9 mix substitution buffer (for test without metabolic activation),
100 µL Bacteria suspension (cf. test system, pre-culture of the strains),
2000 µL Overlay agar
Experiment II (Pre-Incubation)
In the pre-incubation assay 100 µL test solution (solvent or reference mutagen solution (positive control)), 500 µL S9 mix / S9 mix substitution buffer and 100 µL bacterial suspension were mixed in a test tube and incubated at 37°C for 60 minutes. After pre-incubation 2.0 mL overlay agar (45°C) was added to each tube.
The mixture was poured on minimal agar plates. After solidification the plates were incubated upside down for at least 48 hours at 37°C in the dark.
In parallel to each test a sterile control of the test item was performed and documented in the raw data. Therefore, 100 µL of the stock solution, 500 µl S9 mix / S9 mix substitution buffer were mixed with 2.0 mL overlay agar and poured on minimal agar plates.
CONTROLS:
- Vehicle/solvent control: DMSO
- Negative (untreated) controls were performed to assess the spontaneous revertant colony rate.
- Positive control items used demonstrated a direct and indirect acting mutagenic effect depending on the presence or absence of metabolic activation
NUMBER OF REPLICATIONS: Triplicate
OTHER: PRE-EXPERIMENT: To evaluate the toxicity of the test item a pre-experiment was performed with all strains used. Eight concentrations were tested for toxicity and mutation induction with each 3 plates - Rationale for test conditions:
- In the pre-experiment the concentration range of the test item was 3 – 5000 µg/plate. The pre-experiment is reported as experiment I. Since toxic effects were observed in experiment I, a minimum of six concentrations was tested in experiment II. Based on the observed toxic effects 1000 µg/plate were chosen as maximum concentration for strain TA 100. For the remaining strains the maximum concentration was 5000 µg/plate. The concentration range included two logarithmic decades.
The following concentrations were tested in experiment II:
Strain TA 100: 0.3; 1; 3; 10; 33; 100; 333; and 1000 µg/plate
The remaining strains: 10; 33; 100; 333; 1000; 2500; and 5000 µg/plate - Evaluation criteria:
- The Salmonella typhimurium and Escherichia coli reverse mutation assay is considered acceptable if it meets the following criteria:
•regular background growth in the negative and solvent control;
•the spontaneous reversion rates in the negative and solvent control are in the range of our historical data;
• the positive control substances should produce an increase above the threshold of twice (strains TA 98, TA 100, and WP2 uvrA) or thrice (strains TA 1535 and TA 1537) the colony count of the corresponding solvent control;
•a minimum of five analysable dose levels should be present with at least three dose levels showing no signs of toxic effects, evident as a reduction in the number of revertants below the indication factor of 0.5;
•if precipitation occurs a minimum of four concentrations showing no precipitation should be analysed. - Statistics:
- None
Results and discussion
Test resultsopen allclose all
- Key result
- Species / strain:
- S. typhimurium TA 1535
- 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
- Key result
- Species / strain:
- S. typhimurium TA 1537
- 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
- Key result
- Species / strain:
- S. typhimurium TA 98
- Metabolic activation:
- with and without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- cytotoxicity
- Remarks:
- 5000 µg/plate in Exp 1 without S9, 5000 µg/plate in Exp 1 with S9, 2500 to 5000 µg/plate in Exp 2 without S9, 2500 to 5000 µg/plate in Exp 2 with S9,
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- valid
- 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:
- 333 to 5000 µg/plate in Exp 1 without S9, 1000 to 5000 µg/plate in Exp 1 with S9, 100 to 1000 µg/plate in Exp 2 without S9, 333 to 1000 µg/plate in Exp 2 with S9,
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- valid
- Positive controls validity:
- valid
- Key result
- Species / strain:
- E. coli WP2 uvr A
- 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
- Additional information on results:
- Precipitation: Precipitation of the test item in the overlay agar on the incubated agar plates was observed from 2500 to 5000 µg/plate in both experiments. Based on a lower top dose of 1000 µg/plate no precipitation of the test substance was observed on the incubated agar plates in strain TA 100 in experiment II. The undissolved particles had no influence on the data recording.
The plates incubated with the test item showed reduced background growth at the following concentrations (µg/plate):
Strain Experiment I Experiment II
without S9 mix with S9 mix without S9 mix ith S9 mix
TA 1535 / / / /
TA 1537 1000 – 5000 1000 – 5000 / /
TA 98 1000 – 5000 1000 – 5000 1000 – 5000 1000 – 5000
TA 100 333 – 5000 1 000 – 5000 3 33 – 1000 333 – 1000
WP2 uvrA / / / /
MUTAGENICITY
- The vehicle (DMSO) control plates gave counts of revertant colonies within the normal range. All of the positive control chemicals used in the test induced marked increases in the frequency of revertant colonies, both with or without metabolic activation. Thus, the sensitivity of the assay and the efficacy of the S9-mix were validated.
No substantial increase in revertant colony numbers of any of the five tester strains was observed following treatment with Blue chamomilla - European origin at any concentration level, neither in the presence nor absence of metabolic activation (S9 mix). There was also no tendency of higher mutation rates with increasing concentrations in the range below the generally acknowledged border of biological relevance.
Appropriate reference mutagens were used as positive controls. They showed a distinct in-crease in induced revertant colonies.
In experiment II the number of colonies did not quite reach the lower limit of our historical control data in the negative control strain Wp2 uvrA without S9 mix. Since this deviation is rather small, this effect is judged to be based upon statistical fluctuations and has no detrimental impact on the outcome of the study
- Refer Tables 7.6.1/1 and 7.6.1/2 for more details and attached background for individual results.
HISTORICAL CONTROL DATA (with ranges, means and standard deviation
- Positive historical control data: Refer Table 7.6.1/3
- Negative (solvent/vehicle) historical control data: Refer Table 7.6.1/3
Any other information on results incl. tables
Table 7.6.1 /1 Summary of Experiment I
Study Name: 1875200 |
Study Code: Envigo 1875200 |
Experiment: 1875200 VV Plate |
Date Plated: 09.02.2018 |
Assay Conditions: |
Date Counted: 13.02.2018 |
Metabolic Activation |
Test Group |
Dose Level (per plate) |
Revertant Colony Counts (Mean ±SD) |
||||
|
|
|
|
|
|
|
|
|
|
|
TA 1535 |
TA 1537 |
TA 98 |
TA 100 |
WP2 uvrA |
|
|
|
|
|
|
|
|
Without Activation |
DMSO |
|
12 ± 3 |
12 ± 2 |
27 ± 8 |
173 ± 20 |
28 ± 2 |
Untreated |
|
7 ± 3 |
12 ± 3 |
30 ± 8 |
179 ± 10 |
37 ± 11 |
|
Blue |
3 µg |
11 ± 3 |
13 ± 6 |
24 ± 3 |
183 ± 6 |
28 ± 3 |
|
chamomilla- |
10 µg |
12 ± 2 |
9 ± 3 |
23 ± 4 |
180 ± 21 |
29 ± 9 |
|
European |
33 µg |
9 ± 3 |
9 ± 1 |
23 ± 4 |
161 ± 9 |
30 ± 11 |
|
origin |
100 µg |
9 ± 1 |
10 ± 2 |
30 ± 6 |
144 ± 1 |
27 ± 6 |
|
|
333 µg |
12 ± 5 |
14 ± 5 |
24 ± 3 |
47 ± 1R |
32 ± 8 |
|
|
1000 µg |
12 ± 2 |
14 ± 4R |
24 ± 2R |
17 ± 5M R |
32 ± 3 |
|
|
2500 µg |
12 ± 4P M |
14 ± 2P R |
15 ± 3P R |
3 ± 1P M R |
26 ± 8P |
|
|
5000 µg |
8 ± 3P M |
9 ± 1P M R |
7 ± 1P M R |
1 ± 1P M R |
30 ± 8P M |
|
NaN3 |
10 µg |
868 ± 74 |
|
|
2222 ± 194 |
|
|
4-NOPD |
10 µg |
|
|
384 ± 11 |
|
|
|
4-NOPD |
50 µg |
|
59 ± 2 |
|
|
|
|
MMS |
2.0 µL |
|
|
|
|
808 ± 4 |
|
|
|
|
|
|
|
|
|
With Activation |
DMSO |
|
11 ± 2 |
12 ± 1 |
42 ± 8 |
185 ± 9 |
37 ± 10 |
Untreated |
|
12 ± 2 |
12 ± 4 |
49 ± 6 |
189 ± 36 |
44 ± 10 |
|
Blue |
3 µg |
12 ± 3 |
16 ± 1 |
44 ± 6 |
149 ± 3 |
36 ± 4 |
|
chamomilla- |
10 µg |
14 ± 3 |
12 ± 4 |
36 ± 5 |
146 ± 11 |
28 ± 8 |
|
European |
33 µg |
14 ± 3 |
13 ± 3 |
42 ± 9 |
165 ± 12 |
35 ± 11 |
|
origin |
100 µg |
14 ± 3 |
17 ± 3 |
37 ± 2 |
132 ± 16 |
44 ± 10 |
|
|
333 µg |
15 ± 5 |
13 ± 3 |
30 ± 3 |
87 ± 6 |
37 ± 4 |
|
|
1000 µg |
12 ± 3 |
16 ± 1R |
25 ± 8R |
25 ± 6R |
30 ± 9 |
|
|
2500 µg |
11 ± 4P M |
11 ± 3P M R |
30 ± 9P R |
6 ± 2P M R |
29 ± 10P M |
|
|
5000 µg |
6 ± 1P M |
12 ± 3P M R |
18 ± 3P M R |
2 ± 2P M R |
30 ± 8P M |
|
2-AA |
2.5 µg |
506 ± 8 |
143 ± 18 |
3923 ± 200 |
5457 ± 247 |
|
|
2-AA |
10.0 µg |
|
|
|
|
475 ± 38 |
Table 7.6.1/2 Summary of Experiment II
Study Name: 1875200 |
Study Code: Envigo 1875200 |
Experiment: 1875200 HV2 Pre |
Date Plated: 20.02.2018 |
Assay Conditions: |
Date Counted: 27.02.2018 |
Metabolic Activation |
Test Group |
Dose Level (per plate) |
Revertant Colony Counts (Mean ±SD) |
||||
|
|
|
|
|
|
|
|
|
|
|
TA 1535 |
TA 1537 |
TA 98 |
TA 100 |
WP2 uvrA |
|
|
|
|
|
|
|
|
Without Activation |
DMSO |
|
9 ± 3 |
8 ± 2 |
26 ± 10 |
136 ± 17 |
29 ± 2 |
Untreated |
|
10 ± 3 |
8 ± 3 |
29 ± 7 |
196 ± 40 |
28 ± 4 |
|
Blue |
0.3 µg |
|
|
|
139 ± 8 |
|
|
chamomilla- |
1 µg |
|
|
|
125 ± 15 |
|
|
European |
3 µg |
|
|
|
122 ± 7 |
|
|
origin |
10 µg |
|
|
|
109 ± 6 |
|
|
|
33 µg |
9 ± 4 |
7 ± 2 |
27 ± 8 |
84 ± 18 |
23 ± 0 |
|
|
100 µg |
8 ± 4 |
10 ± 2 |
17 ± 4 |
32 ± 3 |
26 ± 4 |
|
|
333 µg |
8 ± 3 |
8 ± 1 |
15 ± 5 |
29 ± 2R |
27 ± 6 |
|
|
1000 µg |
9 ± 3 |
10 ± 1 |
15 ± 2R |
16 ± 2M R |
26 ± 8 |
|
|
2500 µg |
7 ± 2P |
7 ± 3P |
8 ± 0P M R |
|
28 ± 2P |
|
|
5000 µg |
9 ± 1P M |
5 ± 1P M |
2 ± 1P M R |
|
26 ± 7P M |
|
NaN3 |
10 µg |
1145 ± 70 |
|
|
2142 ± 148 |
|
|
4-NOPD |
10 µg |
|
|
370 ± 30 |
|
|
|
4-NOPD |
50 µg |
|
52 ± 15 |
|
|
|
|
MMS |
2.0 µL |
|
|
|
|
680 ± 56 |
|
|
|
|
|
|
|
|
|
With Activation |
DMSO |
|
13 ± 4 |
12 ± 2 |
35 ± 9 |
107 ± 8 |
39 ± 12 |
Untreated |
|
13 ± 4 |
13 ± 2 |
41 ± 4 |
177 ± 7 |
32 ± 1 |
|
Blue |
0.3 µg |
|
|
|
108 ± 3 |
|
|
chamomilla- |
1 µg |
|
|
|
110 ± 14 |
|
|
European |
3 µg |
|
|
|
124 ± 13 |
|
|
origin |
10 µg |
|
|
|
124 ± 17 |
|
|
|
33 µg |
13 ± 6 |
13 ± 4 |
34 ± 7 |
113 ± 6 |
43 ± 10 |
|
|
100 µg |
13 ± 1 |
9 ± 3 |
31 ± 6 |
118 ± 6 |
41 ± 8 |
|
|
333 µg |
12 ± 4 |
12 ± 1 |
24 ± 1 |
39 ± 5R |
37 ± 11 |
|
|
1000 µg |
12 ± 2 |
13 ± 3 |
26 ± 9R |
23 ± 12R |
32 ± 10 |
|
|
2500 µg |
12 ± 4P |
12 ± 4P M |
15 ± 4P M R |
|
25 ± 8P M |
|
|
5000 µg |
8 ± 2P |
14 ± 3P M |
3 ± 2P M R |
|
27 ± 5P M |
|
2-AA |
2.5 µg |
365 ± 7 |
197 ± 14 |
4121 ± 575 |
4029 ± 522 |
|
|
2-AA |
10.0 µg |
|
|
|
|
245 ± 33 |
|
|
|
|
|
|
|
|
|
Key to Positive Controls |
Key to Plate Postfix Codes |
||
|
|
||
NaN3 2-AA 4-NOPD MMS |
sodium azide 2-aminoanthracene 4-nitro-o-phenylene-diamine methyl methane sulfonate |
R M P |
Reduced background growth Manual count Precipitate |
Table 7.6.1/3/ Historical data
Strain |
|
without S9 mix |
with S9 mix |
||||||
|
|
Mean |
SD |
Min |
Max |
Mean |
SD |
Min |
Max |
|
Solvent control |
12 |
2.5 |
6 |
25 |
12 |
2.5 |
7 |
26 |
TA 1535 |
Untreated control |
12 |
3.1 |
6 |
28 |
12 |
2.9 |
7 |
26 |
|
Positive control |
1130 |
143.1 |
334 |
1816 |
388 |
58.2 |
176 |
668 |
|
Solvent control |
10 |
2.2 |
6 |
19 |
13 |
3.5 |
7 |
30 |
TA1537 |
Untreated control |
11 |
2.7 |
5 |
21 |
14 |
4.0 |
7 |
31 |
|
Positive control |
82 |
12.7 |
43 |
157 |
191 |
60.8 |
83 |
434 |
|
Solvent control |
25 |
4.4 |
13 |
43 |
34 |
6.2 |
15 |
58 |
TA 98 |
Untreated control |
27 |
4.9 |
12 |
43 |
37 |
6.5 |
11 |
57 |
|
Positive control |
378 |
73.7 |
211 |
627 |
3949 |
771.8 |
360 |
6586 |
|
Solvent control |
156 |
26.0 |
78 |
209 |
148 |
32.3 |
73 |
208 |
TA 100 |
Untreated control |
176 |
23.6 |
79 |
217 |
172 |
25.4 |
85 |
218 |
|
Positive control |
1966 |
293.2 |
498 |
2767 |
3798 |
830.4 |
536 |
6076 |
|
Solvent control |
41 |
5.6 |
27 |
63 |
50 |
6.8 |
28 |
72 |
WP2uvrA |
Untreated control |
42 |
5.8 |
30 |
63 |
52 |
6.8 |
36 |
88 |
|
Positive control |
798 |
362.7 |
319 |
4732 |
378 |
112.6 |
167 |
1265 |
Mean = mean value of revertants/plate
SD = standard deviation
Min = minimal value/Max = maximal value
Applicant's summary and conclusion
- Conclusions:
- In conclusion, it can be stated that during the described mutagenicity test and under the experimental conditions reported, the test item did not induce gene mutations by base pair changes or frameshifts in the genome of the strains used.
- Executive summary:
This study was performed to investigate the potential of Blue chamomilla - European origin to induce gene mutations according to the plate incorporation test (experiment I) and the pre-incubation test (experiment II) using theSalmonella typhimuriumstrains TA 1535, TA 1537, TA 98, TA 100, and the Escherichia colistrain WP2 uvrA.
The assay was performed in two independent experiments both with and without liver microsomal activation. Each concentration, including the controls, was tested in triplicate. The test item was tested at the following concentrations:
Pre-Experiment/Experiment I: 3; 10; 33; 100; 333; 1000; 2500; and 5000 µg/plate
Experiment II:
Strains TA 100: 0.3; 1; 3; 10; 33; 100; 333; and 1000 µg/plate
The remaining strains: 33; 100; 333; 1000; 2500; and 5000 µg/plate
The test item precipitated in the overlay agar in the test tubes from 1000 to 5000 µg/plate in experiment I and from 333 µg/plate up to the highest investigated dose in experiment II. Precipitation of the test item in the overlay agar on the incubated agar plates was observed from 2500 to 5000 µg/plate in both experiments. Based on a lower top dose of 1000 µg/plate no precipitation of the test substance was observed on the incubated agar plates in strain TA 100 in experiment II. The undissolved particles had no influence on the data recording.
The plates incubated with the test item showed reduced background in strains TA 1537, TA 98, and TA 100.
Toxic effects, evident as a reduction in the number of revertants (below the indication factor of 0.5), occurred in strains TA 98 and TA 100 with and without S9 mix in both experiments .
No substantial increase in revertant colony numbers of any of the fivetester strains was observed following treatment with Blue chamomilla - European origin at any dose level, neither in the presence nor absence of metabolic activation (S9 mix). There was also no tendency of higher mutation rates with increasing concentrations in the range below the generally acknowledged border of biological relevance.
Appropriate reference mutagens were used as positive controls and showed a distinct increase of induced revertant colonies.
In conclusion, it can be stated that during the described mutagenicity test and under the experimental conditions reported, the test item did not induce gene mutations by base pair changes or frameshifts in the genome of the strains used.
Therefore, Blue chamomilla - European origin is considered to be non-mutagenic in thisSalmonella typhimuriumandEscherichia colireverse mutation assay.
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