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EC number: - | 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
Skin sensitisation
Administrative data
- Endpoint:
- skin sensitisation: in vivo (LLNA)
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Study period:
- The study was performed between 28 August 2012 and 11 September 2012.
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- other: Study conducted to GLP and in compliance with agreed protocols, with no or minor deviations from standard test guidelines and/or minor methodological deficiencies, which do not effect the quality of the relevant results.
Data source
Reference
- Reference Type:
- study report
- Title:
- Unnamed
- Year:
- 2 012
Materials and methods
Test guidelineopen allclose all
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)
- Deviations:
- no
- Qualifier:
- according to guideline
- Guideline:
- EU Method B.42 (Skin Sensitisation: Local Lymph Node Assay)
- Deviations:
- no
- GLP compliance:
- yes (incl. QA statement)
- Type of study:
- mouse local lymph node assay (LLNA)
Test material
- Test material form:
- other: solid
Constituent 1
In vivo test system
Test animals
- Species:
- mouse
- Strain:
- CBA
- Sex:
- female
- Details on test animals and environmental conditions:
- TEST ANIMALS
- Source: Female CBA/Ca (CBA/CaOlaHsd) strain mice were supplied by Harlan Laboratories UK Ltd.
- Age at study initiation: eight to twelve weeks old
- Weight at study initiation: 15 to 23 g
- Housing: The animals were individually housed in suspended solid floor polypropylene cages furnished with softwood woodflakes.
- Diet (e.g. ad libitum): Free access to food (2014C Teklad Global Rodent diet) was allowed throughout the study.
- Water (e.g. ad libitum): Free access to mains tap water was allowed throughout the study.
- Acclimation period: at least five days
ENVIRONMENTAL CONDITIONS
- Temperature (°C): The temperature was controlled to remain within target range of 19 to 25°C
- Humidity (%): The relative humidity was controlled to remain within target range of 30 to 70%,
- Air changes (per hr): The rate of air exchange was approximately fifteen changes per hour.
- Photoperiod (hrs dark / hrs light): Lighting was controlled by a time switch to give twelve hours continuous light (06.00 to 18.00) and twelve hours darkness.
Study design: in vivo (LLNA)
- Vehicle:
- dimethylformamide
- Concentration:
- Preliminary Screening Test: 25%
Main Test: 25%, 10% and 5% - No. of animals per dose:
- Preliminary Screening Test: 1 mouse
Main Test: Five mice per dose - Details on study design:
- PREPARATION OF TEST ITEM:
For the purpose of the study, the test item was freshly prepared as a suspension in dimethyl formamide. This vehicle was chosen as it produced the highest concentration that was suitable for dosing.
The test item was formulated within two hours of being applied to the test system. It is assumed that the formulation was stable for this duration.
PRELIMINARY SCREENING TEST:
Using available information regarding the systemic toxicity/irritancy potential of the test item, a preliminary screening test was performed using one mouse. The mouse was treated by daily application of 25 µl of the test item at a maximum attainable concentration of 25% w/w in dimethyl formamide, to the dorsal surface of each ear for three consecutive days (Days 1, 2, 3). The mouse was observed twice daily on Days 1, 2 and 3 and once daily on Days 4, 5 and 6. Local irritation was scored daily according to the scale for erythema below. Any clinical signs of toxicity, if present, were also recorded. The bodyweight was recorded on Day 1 (prior to dosing) and on Day 6.
Scale for Erythema :
Observation
No erythema: 0
Very slight erythema (barely perceptible): 1
Well-defined erythema: 2
Moderate to severe erythema: 3
Severe erythema (beef redness) to eschar formation preventing grading of erythema: 4
The thickness of each ear was measured using an Oditest micrometer, pre dose on Day 1, post dose on Day 3 and on Day 6. Any changes in the ear thickness were noted. Mean ear thickness changes were calculated between Days 1 to 3 and Days 1 to 6. A mean ear thickness increase of equal to or greater than 25% was considered to indicate excessive irritation and limited biological relevance to the endpoint of sensitisation.
MAIN TEST:
TEST ITEM ADMINISTRATION:
Groups of five mice were treated with the test item at concentrations of 25%, 10% or 5% w/w in dimethyl formamide. The preliminary screening test suggested that the test item would not produce systemic toxicity or excessive local irritation at the highest suitable concentration. The mice were treated by daily application of 25 µl of the appropriate concentration of the test item to the dorsal surface of each ear for three consecutive days (Days 1, 2, 3). The test item formulation was administered using an automatic micropipette and spread over the dorsal surface of the ear using the tip of the pipette.
A further group of five mice received the vehicle alone in the same manner.
The positive control animals were similarly treated to the test animals except that 25 µl of the positive control item, α Hexylcinnamaldehyde, tech., 85%, at a concentration of 15% v/v in dimethyl formamide was applied to the dorsal surface of each ear.
The thickness of each ear was measured using an Oditest micrometer, pre dose on Day 1 and on Days 2, 3, 4, 5 and 6. Any changes in the ear thickness were noted. Daily mean ear thickness changes were calculated. A mean ear thickness increase of equal to or greater than 25% was considered to indicate excessive irritation and limited biological relevance to the endpoint of sensitisation.
3H-Methyl Thymidine Administration:
Five days following the first topical application of the test item, vehicle control or positive control item (Day 6) all mice were injected via the tail vein with 250 µl of phosphate buffered saline (PBS) containing 3H methyl thymidine (3HTdR:80µCi/ml, specific activity 2.0 Ci/mmol,) giving a total of 20 µCi to each mouse.
OBSERVATIONS:
Clinical Observations: All animals were observed twice daily on Days 1, 2 and 3 and on a daily basis on Days 4, 5 and 6. Any signs of toxicity or signs of ill health during the test were recorded.
Local Skin Irritation Observations: All animals were observed daily. Any signs of local skin irritation noted during the test were recorded.
Bodyweights: The bodyweight of each mouse was recorded on Day 1 (prior to dosing) and Day 6 (prior to termination).
TERMINAL PROCEDURES:
Termination: Five hours following the administration of 3HTdR all mice were killed by carbon dioxide asphyxiation followed by cervical separation. For each individual animal of each group the draining auricular lymph nodes were excised and processed. For each individual animal 1 ml of PBS was added to the lymph nodes.
Preparation of Single Cell Suspension: A single cell suspension of the lymph node cells for each individual animal was prepared by gentle mechanical disaggregation through a 200 mesh stainless steel gauze. The lymph node cells were rinsed through the gauze with 4 ml of PBS into a petri dish labelled with the project number and dose concentration. The lymph node cells suspension was transferred to a centrifuge tube. The petri dish was washed with an additional 5 ml of PBS to remove all remaining lymph node cells and these were added to the centrifuge tube. The lymph node cells were pelleted at 1400 rpm (approximately 190 g) for ten minutes. The pellet was resuspended in 10 ml of PBS and re-pelleted. To precipitate out the radioactive material, the pellet was resuspended in 3 ml of 5% Trichloroacetic acid (TCA).
Determination of 3HTdR Incorporation: After approximately eighteen hours incubation at approximately 4°C, the precipitates were recovered by centrifugation at 2100 rpm (approximately 450 g) for ten minutes, resuspended in 1 ml of TCA and transferred to 10 ml of scintillation fluid (Optiphase 'Trisafe'). 3HTdR incorporation was measured by beta-scintillation counting. The "Poly QTM" vials containing the samples and scintillation fluid were placed in the sample changer of the scintillator and left for approximately twenty minutes. The purpose of this period of time in darkness was to reduce the risk of luminescence, which has been shown to affect the reliability of the results. After approximately twenty minutes, the vials were shaken vigorously. The number of radioactive disintegrations per minute was then measured using the Beckman LS6500 scintillation system. - Positive control substance(s):
- other: α-Hexylcinnamaldehyde, tech., 85%. The positive control item was freshly prepared as a 15% v/v dilution in dimethyl formamide.
- Statistics:
- Data was processed to give group mean values for disintegrations per minute and standard deviations where appropriate. Individual and group mean disintegrations per minute values were assessed for dose response relationships by analysis of homogeneity of variance followed by one way analysis of variance (ANOVA). In the event of a significant result from the ANOVA, pairwise comparisons were performed between control and treated groups. For homogenous datasets Dunnett’s Multiple Comparison test was used and for non homogenous datasets Dunnett’s T3 Multiple Comparison Method was used.
Probability values (p) are presented as follows:
P<0.001 ***
P<0.01 **
P<0.05 *
P>0.05 (not significant)
Results and discussion
- Positive control results:
- The positive control item, α-Hexylcinnamaldehyde, tech., 85%, gave a Stimulation Index of greater than 3 when tested at a concentration of 15% w/w in dimethyl formamide.
In vivo (LLNA)
Resultsopen allclose all
- Parameter:
- SI
- Value:
- 0.81
- Test group / Remarks:
- Test item 5% w/w in dimethyl formamide
- Parameter:
- SI
- Value:
- 1.51
- Test group / Remarks:
- Test item 10% w/w in dimethyl formamide
- Parameter:
- SI
- Value:
- 1.24
- Test group / Remarks:
- Test item 25% w/win dimethyl formamide
Any other information on results incl. tables
Preliminary Screening Test
Clinical observations, bodyweight and mortality data are given in Table 1 and local skin irritation is given in Table 2. The ear thickness measurements and mean ear thickness changes are given in Table 3 (attached background material).
Off white residual test item on the ears was noted on Days 1 to 5. No signs of systemic toxicity,visual local skin irritation or excessive irritation indicated by anequal to or greater than 25% increase in mean ear thickness were noted.
Based on this information the dose levels selected for the main test were 25%,10% and 5% w/w in dimethyl formamide.
Main Test
Estimation of the Proliferative Response of Lymph Node Cells
The radioactive disintegrations per minute per animal and the stimulation index are given in Table 4 (attached background material).
The Stimulation Index expressed as the mean radioactive incorporation for each treatment group divided by the mean radioactive incorporation of the vehicle control group are as follows:
Treatment Group |
Concentration |
Stimulation Index |
Result |
Test Item |
5% w/w in |
0.81 |
Negative |
10% w/w in |
1.51 |
Negative |
|
25% w/w in |
1.24 |
Negative |
|
Positive |
15% v/v in |
5.19 |
Positive |
Clinical Observations and Mortality Data
Individual clinical observations and mortality data for test and control animals are given in Table 5 and local skin irritation is given in Table 6. The ear thickness measurements and mean ear thickness changes are given in Table 7 (see attached background material).
Off white residual test item on the ears was noted in animals treated with the test item.
There were no deaths. No signs of systemic toxicity or excessive irritation indicated by anequal to or greater than 25% increase in mean ear thickness were noted in the test or control group animals during the test. No visual local skin irritation was noted in animals treated with the test item or vehicle control item during the test. Very slight erythema was noted on Days 2 and 3 on both ears of the animals treated with the positive control item.
Bodyweight:
Individual bodyweights and bodyweight changes for test and control animals are given in Table 8 (attached background material).
Bodyweight changes of the test animals between Day 1 and Day 6 were comparable to those observed in the corresponding control group animals over the same period.
Applicant's summary and conclusion
- Interpretation of results:
- not sensitising
- Remarks:
- Migrated information Criteria used for interpretation of results: EU
- Conclusions:
- The test item was considered to be a non-sensitiser under the conditions of the test.
The test item did not meet the criteria for classification according to the Globally Harmonised System of Classification and Labelling of Chemicals.
The positive control item, α-Hexylcinnamaldehyde, tech., 85%, gave a Stimulation Index of greater than 3 when tested at a concentration of 15% w/w in dimethyl formamide - Executive summary:
Introduction
A study was performed to assess the skin sensitisation potential of the test item in the CBA/Ca strain mouse following topical application to the dorsal surface of the ear. The method was designed tobe compatible with the following:
-OECD Guideline for the Testing of Chemicals No. 429 "Skin Sensitisation: Local Lymph Node Assay" (adopted 22 July 2010)
- Method B42 Skin Sensitisation (Local Lymph Node Assay) of Commission Regulation (EC) No. 440/2008
Methods
Following a preliminary screening test in which no clinical signs of toxicity were noted at a maximum attainable concentration of 25% w/w, this concentration was selected as the highest dose investigated in the main test of the Local Lymph Node Assay. Three groups, each of five animals, were treated with 50 µl (25 µl per ear) of the test item as a suspension in dimethyl formamide at concentrations of 25%,10% or 5% w/w. A further group of five animals was treated with dimethyl formamide alone. A concurrent positive control test, using a group of five animals, was also performed with the known sensitiser, α‑Hexylcinnamaldehyde tech., 85%, at a concentration of 15% v/v in dimethyl formamide.
Results
The Stimulation Index expressed as the mean radioactive incorporation for each treatment group divided by the mean radioactive incorporation of the vehicle control group are as follows:
Treatment Group
Concentration
Stimulation Index
Result
Test Item
5% w/w in
dimethyl formamide0.81
Negative
10% w/w in
dimethyl formamide1.51
Negative
25% w/w in
dimethyl formamide1.24
Negative
Positive
Control Item15% v/v in
dimethyl formamide5.19
Positive
Conclusion
The test item was considered to be a non-sensitiserunder the conditions of the test.
The test item did not meet the criteria for classification according to the Globally Harmonised System of Classification and Labelling of Chemicals.
The positive control item, α-Hexylcinnamaldehyde, tech., 85%, gave a Stimulation Index of greater than 3 when tested at a concentration of 15% w/wi n dimethyl formamide.
Information on Registered Substances comes from registration dossiers which have been assigned a registration number. The assignment of a registration number does however not guarantee that the information in the dossier is correct or that the dossier is compliant with Regulation (EC) No 1907/2006 (the REACH Regulation). This information has not been reviewed or verified by the Agency or any other authority. The content is subject to change without prior notice.
Reproduction or further distribution of this information may be subject to copyright protection. Use of the information without obtaining the permission from the owner(s) of the respective information might violate the rights of the owner.
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