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Toxicological information

Skin sensitisation

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Administrative data

Endpoint:
skin sensitisation: in vivo (LLNA)
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: OECD guideline study, conducted under GLP.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2010
Report Date:
2010

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.42 (Skin Sensitisation: Local Lymph Node Assay)
Deviations:
no
Qualifier:
according to
Guideline:
EPA OPPTS 870.2600 (Skin Sensitisation)
Deviations:
no
GLP compliance:
yes
Type of study:
mouse local lymph node assay (LLNA)

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
Identification: tetrachloroethyleneTrade name (Dow): DOWPER MCDescription: clear colourless liquidBatch nr: 1025168Purity: Perchloroethylene 99.6%

In vivo test system

Test animals

Species:
mouse
Strain:
other: CBA/J strain, inbred, SPF quality
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS- Source: Female mice, CBA/J strain, inbred were supplied by Harlan, Horst, The Netherlands- Age at study initiation: approx. 10 weeks old- Weight at study initiation: 19-24 grams- Housing: individual housing in labeled Macrolon cages (MI type; height 12.5 cm) containing sterilized sawdust as bedding material.- Diet (e.g. ad libitum): free access to pelleted rodent diet - Water (e.g. ad libitum): Free access to tap water- Acclimation period: at least 5 days before the start of the treatment, under laboratory conditons.ENVIRONMENTAL CONDITIONS- Temperature (°C): 21.0 ± 3.0ºC (actual range: 19.7 - 23.1ºC),- Humidity (%): 40-70% (actual range: 31 - 65%) - Air changes (per hr): approximately 15 air changes per hour- Photoperiod (hrs dark / hrs light): 12 hours artificial fluorescent light and 12 hours darkness per day.IN-LIFE DATES: From: day 1 To: Day 6

Study design: in vivo (LLNA)

Vehicle:
acetone/olive oil (4:1 v/v)
Concentration:
In the main study, three experimental groups of five female CBA/J mice were treated with test substance concentrations of 5, 25 or 100% w/w on three consecutive days, by open application on the ears. Five vehicle control animals were similarly treated, but with vehicle alone (Acetone/Olive oil (4:1 v/v)).
No. of animals per dose:
Groups of five mice were treated.
Details on study design:
RANGE FINDING TESTS:Two test material concentrations were tested; a 50% and 100% concentration. The highest concentration was the maximum concentration as required in the test guidelines (undiluted for liquids). The test system, procedures and techniques were identical to those used during Days 1 to 3 of the main study. Two young adult animals were selected (8-14 weeks old). Each animal was treated with one concentration on three consecutive days. Approximately 3-4 hours after the last exposure, the irritation of the ears was assessed. Body weights were determined on Day 3. The animals were sacrificed after the final observation and no necropsy was performed.MAIN STUDYANIMAL ASSIGNMENT AND TREATMENT- Name of test method: Local Lymph Node Assay in the mouse. - Criteria used to consider a positive response: Disintegrations Per Minute (DPM) values are presented for each animal and for each dose group. A Stimulation Index (SI) is calculated for each group. The SI is the ratio of the DPM/group compared to DPM/vehicle control group. If the results indicate a SI ≥ 3, the test material may be regarded as a skin sensitizer, based on the test guideline and recommendations done by ICCVAM. The results were evaluated according to the Globally Harmonized System of ClassificationTREATMENT PREPARATION AND ADMINISTRATION:Induction - Days 1, 2 and 3The highest test substance concentration (100%) used in the main study was selected from the preliminary irritation study. Three experimental groups of five female CBA/J mice were treated with test substance concentrations of 5, 25 or 100% w/w on three consecutive days. The dorsal surface of both ears was epidermally treated (25 μL/ear) with the test substance concentration, at approximately the same time per day. The concentrations were mixed thoroughly using a vortex mixer immediately prior to dosing. The control animals were treated the same as the experimental animals, except that, instead of the test substance, the vehicle alone was administered. Excision of the nodes - Day 6Three days after the last exposure, all animals were injected with 3H-methyl thymidine and after five hours the draining (auricular) lymph nodes wereexcised and pooled for each animal. After precipitating the DNA of the lymph node cells, radioactivity measurements were performed. The activity was expressed as the number of Disintegrations Per Minute (DPM) and a stimulation index (SI) was subsequently calculated for each group.
Positive control substance(s):
hexyl cinnamic aldehyde (CAS No 101-86-0)

Results and discussion

Positive control results:
A reliability check is carried out at regular intervals to check the sensitivity of the test system and the reliability of the experimental techniques as used by NOTOX. In this study, performed in September/October 2009, females of the CBA/J mouse strain (Charles River France, L’Arbresle Cedex, France) were checked for sensitivity to Alpha- Hexylcinnamaldehyde, technical grade. The females were approx. 11 weeks old at commencement of the study. The study was based on the OECD Guideline No. 429, EC No 440/2008, Part B.42 and EPA, OPPTS 870.2600 “Skin Sensitization”. Alpha-hexylcinnamicaldehyde, technical grade (CAS no. 101-86-0) was fabricated under lot no. 13102MO (Sigma- Aldrich, Steinheim, Germany) and the 3H-methylthymidine was purchased from PerkinElmer Life Sciences, Boston, MA, USA. HCA concentrations used for this study were 5, 10 and 25% in Acetone/Olive oil (4:1 (v/v)). The SI values calculated for the substance concentrations 5, 10 and 25% were 1.4, 1.2 and 5.1 respectively. An EC3 value of 16.9% was calculated using linear interpolation. The calculated EC3 value was found to be in the acceptable range of 2 and 20%. The results of the 6 monthly HCA reliability checks of the recent years were 13.1, 15.6, 14.1, 13.8, 13.9, 16.0 and 11.9%. Based on the results, it was concluded that the Local Lymph Node Assay as performed at NOTOX is an appropriate model for testing for contact hypersensitivity.

In vivo (LLNA)

Resultsopen allclose all
Key result
Parameter:
SI
Remarks on result:
other: The mean ± SEM SI values for the vehicle control and the treated groups (5%, 25% and 100%) were 1.0 ± 0.3, 0.9 ± 0.3, 1.4 ± 0.4 and 4.3 ± 1.4, respectively.
Key result
Parameter:
other: disintegrations per minute (DPM)
Remarks on result:
other: The mean ± SEM DPM values for the vehicle control and the treated groups (5%, 25% and 100%) were 622 ± 116, 533 ± 157, 852 ± 194 and 2690 ± 693, respectively.

Any other information on results incl. tables

See attached PDF with tables 3, 4 and figure 1 from the original report.

Applicant's summary and conclusion

Interpretation of results:
sensitising
Remarks:
Migrated information weak
Conclusions:
The SI values calculated for the test material concentrations 5, 25 and 100% were 0.9, 1.4 and 4.3 respectively. These results indicate that the test material could elicit an SI ≥ 3. An EC3 value (the estimated test material concentration that will give a SI =3) of 66.4% was calculated. Based on these results and according to the recommendations made in the test guidelines, the test material would be regarded as a weak skin sensitiser.Tetrachloroethylene is unlikely to be a sensitizer based the mode of action for small molecules, which would include covalent binding to bio-molecules (proteins). The test material contains a stabiliser at 0.4% classified as a weak skin sensitizer, the uptake of which may be increased by tetrachloroethylene.According to the Regulation (EC) No 1272/2008 on classification, labeling and packaging of substances and mixtures, the test material should be classified as skin sensitizer (Category 1) and labeled as H317: May cause an allergic skin reaction. Based on the EC3 value of 66%, tetrachloroethylene can be categorized as weak sensitizer based on the proposal of ECETOC (European Centre for Ecotoxicology and Toxicology of Chemicals). Contact Sensitisation: Classification According to Potency. Technical Report No. 87. Brussels, April 2003 (ISSN-0773-8072-87).
Executive summary:

The study was carried out according to OECD guideline No. 429. Test substance concentrations selected for the main study were based on the results of a preliminary study. In the main study, three groups of five female CBA/J mice were treated with test substance concentrations of 5, 25 or 100% w/w on three consecutive days, by open application on the ears. Five vehicle control animals were similarly treated, but with vehicle alone (Acetone/Olive oil (4:1 v/v)). Three days after the last exposure, all animals were injected with 3H-methyl thymidine and after five hours the draining (auricular) lymph nodes were excised and pooled for each animal. After precipitating the DNA of the lymph node cells, radioactivity measurements were performed. The activity was expressed as the number of Disintegrations Per Minute (DPM) and a stimulation index (SI) was subsequently calculated for each group. The slight irritation of the ears as shown by all animals treated at 100% and one animal treated at 25% was considered not to have a toxicologically significant effect on the activity of the nodes. The majority of auricular lymph nodes were considered normal in size, except for the nodes in the animals of the highest dose groups. Mean DPM/animal values for the experimental groups treated with test substance concentrations 5, 25 and 100% were 533, 852 and 2690 DPM respectively. The mean DPM/animal value for the vehicle control group was 622 DPM. The SI values calculated for the substance concentrations 5, 25 and 100% were 0.9, 1.4 and 4.3 respectively. These results indicate that the test substance could elicit an SI ≥ 3. An EC3 value (the estimated test substance concentration that will give a SI =3) of 66.4% was calculated. The six monthly reliability check with Hexylcinnamaldehyde indicates that the Local Lymph Node Assay as performed is an appropriate model for testing for contact hypersensitivity.

Based on these results, the test material is regarded as a weak skinsensitizer.Tetrachloroethyleneis unlikely to be a sensitizer based the mode of action for small molecules, which would include covalent binding to bio-molecules (proteins). The test material containsa stabiliserat 0.4% classified as a weak skin sensitizer, the uptake of which may be increased by tetrachloroethylene.