Methyl phenyl sulphide

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

11th May 2010 to 10th August 2010.

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Proprietary guideline study, compliant with GLP.

Cross-reference

Cross-reference 1

Reason / purpose for cross-reference:

reference to same study

Cross-reference 2

Reason / purpose for cross-reference:

reference to other study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2010

Report date:

2010

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)

Deviations:

yes

Remarks:

A deviation from maximum and minimum relative humidity occurred. However, this did not affect the validity of the study.

Principles of method if other than guideline:

Not relevant

GLP compliance:

yes

Type of study:

mouse local lymph node assay (LLNA)

Test material

Test material information

Constituent 1

Reference substance name:

Thioanisole

IUPAC Name:

Thioanisole

Constituent 2

Reference substance name:

Methyl phenyl sulphide

EC Number:

202-878-2

EC Name:

Methyl phenyl sulphide

Cas Number:

100-68-5

Molecular formula:

C7H8S

IUPAC Name:

(methylsulfanyl)benzene

Details on test material:

- Name of test material (as cited in study report): Thioanisole
- Molecular formula (if other than submission substance): C7H8S
- Molecular weight (if other than submission substance): 124.19
- Smiles notation (if other than submission substance): Not documented
- InChl (if other than submission substance): Not documented
- Structural formula attached as image file (if other than submission substance): see Fig. Not documented
- Substance type: Not documented
- Physical state: Colourless to slightly yellowish liquid
- Analytical purity: 99% minimum
- Impurities (identity and concentrations): Not documented
- Composition of test material, percentage of components: Not documented
- Isomers composition: Not documented
- Purity test date: Not documented
- Lot/batch No.: 100310
- Expiration date of the lot/batch: 23rd March 2011
- Radiochemical purity (if radiolabelling): Not documented
- Specific activity (if radiolabelling): Not documented
- Locations of the label (if radiolabelling): Not documented
- Expiration date of radiochemical substance (if radiolabelling): Not documented
- Stability under test conditions: Stable
- Storage condition of test material: At room temperature protected from light under nitrogen.
- Other: Use amber coloured glassware or wrap container in tin foil when handling.

In vivo test system

Test animals

Species:

mouse

Strain:

other: CBA/J strain

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: Harlan, Horst, The Netherlands
- Age at study initiation: approximately 9 weeks old
- Weight at study initiation: 19 - 23g
- Housing: Individually housed in labelled Macrolon cages (MI type; height 12,5cm) containing sterilized sawdust as bedding material. Paper was supplied as cage enrichment. The paper was removed on Day 1 prior to dosing and was supplied again after scoring of the ears on Day 3. During the acclimatization period, the animals were group housed in Macrolon cages (MI type; height 18cm).
- 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 treatment under laboratory conditions.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21.0 ± 3°C (actual range: 19.7 - 22.9°C)
- Humidity (%): 40 - 70% (actual range: 38 - 81%)
- Air changes (per hr): Not documented
- Photoperiod (hrs dark / hrs light): 12 hours artificial fluorescent light and 12 hours darkness per day.

IN-LIFE DATES: From: 17 May 2010 To: 14 June 2010

Study design: in vivo (LLNA)

Vehicle:

acetone/olive oil (4:1 v/v)

Concentration:

0, 10, 25 and 50%

No. of animals per dose:

Five females per group

Details on study design:

RANGE FINDING TESTS:
A preliminary irritation study was conducted in order to select the highest test substance concentration to be used in the main study. Initially two test substance concentrations were tested, 50% and 100% concentration. Two young adult animals were selected. Each animal was treated with one concentration on three consecutive days. Approximately 3-4 days after the last exposure, the irritation of the ears was assessed. Bodyweights were determined on Day 3. The animals were sacrificed after the final observation. No necropsy was performed. Based on the death of the animal at 100% on Day 2, two additional animals were treated in a similar manner with 2 lower concentrations, specifically 25% and 10% immediately after the animals were found dead.

MAIN STUDY
ANIMAL ASSIGNMENT AND TREATMENT
Three groups of five animals were treated with one test substance concentration per group. The highest test substance concentration was selected from the preliminary irritation study. One group of five animals was treated with the vehicle.
- Name of test method: Local Lymph Node Assay
- Criteria used to consider a positive response: If the results indicate a Stimulation Index equal to or greater than 3, the test substance should be considered to be a skin sensitiser.

TREATMENT PREPARATION AND ADMINISTRATION:
The test substance formulations were prepared within 4 hours prior to each treatment. Test substance and formulations were protected from light using amber coloured glassware or aluminium foil. No adjustment was made for specific gravity of the vehicle. Homogeneity was obtained to visually acceptable levels.

Induction:
The dorsal surface of both ears was epidermally treated (25 µL/ear) with the test substance concentration at approximately the same time each day. The concentrations were mixed thoroughly using a vortex mixer immediately prior to dosing. This process occurred for three successive days. On the 6th day, excision of the nodes and processing the tissues for radioactivity occurred. Radioactivity measurements were made on day 7.

Positive control substance(s):

other: Alpha-hexylcinnamicaldehyde

Statistics:

No information provided

Results and discussion

Positive control results:

A six monthly check for assay validity and method integrity was completed using the positive control material alpha- hexylcinnamicaldehyde. This confirmed the methods used and the lab conduct of the assay were acceptable to identify a known weak to moderate sensitiser .

In vivo (LLNA)

Results

Results 1

Parameter:

SI

Remarks on result:

other: The Stimulation Index values calculated for the substance concentrations 10, 25 and 50% were 1, 2.3 and 4.9 respectively. A linear dose response and an SI of greaterthan 3.0 confirmed Thioanisole is a potential skin sensitiser

Results 2

Parameter:

other: disintegrations per minute (DPM)

Remarks on result:

other: The mean DPM/animals values for the experimental groups treated with the test substance concentrations of 10, 25 and 50% were 693, 1508 and 3275 DPM respectively. The mean DPM value for the control group was 662.

Any other information on results incl. tables

Preliminary Study:

Based on the results of this study, the highest test susbtance concentration selected for the main study was a 50% concentration.

Main Study:

Skin reactions:

Slight irritation of the ears was shown by all experimental animals at all three concentrations. However, this was not considered to have a toxicologically significant effect on the activity of the nodes. No oedema was observed in any of the animals examined.

Macroscopic examination:

All auricular lymph nodes of the animals at 50% concentration were considered increased in size. All other auricular lymph nodes were considered normal in size. No macroscopic abnormalities of the surrounding area were noted in any of the animals.

Body Weights:

Body weights and body weight gain of experimental animals remained in the same range as the controls over the study period. The slight body weight loss in some animals was not considered toxicologically significant.

Toxicity / Mortality:

No mortality occurred and no symptoms of systemic toxicity were observed in the animals of the main study.

Applicant's summary and conclusion

Interpretation of results:

sensitising

Remarks:

Migrated information

Conclusions:

Under the conditions of this study, the test substance, Thioanisole, should be considered to be a skin sensitiser. Based on these results, the test substance should be classified as a Category 1 skin sensitizer and have the hazard statement H317: May casue an allergic reaction associated with it, in accordance with Regulation EC No. 1272/2008. According to Directive 67/548/EEC, the test substance should be classified with the risk phrase R43: May cause sensitization by skin contact and have the symbol Xi, Irritant associated with it.

Executive summary:

In a study conducted by Beerens-Heijnen (2010), the test substance, Thioanisole, was examined for its ability to cause skin sensitization when applied to female CBA/J strain of mice in a Local Lymph Node Assay. The test substance was applied to the dorsal surface of the ear of each rabbit for three consecutive days at concentrations of 10, 25 and 50% (w/w). On day 6 of experimentation, the auricular lymph nodes were excised and pooled for each animal. After precipitating the DNA of the lymph node cells, radioactivity measurements were made and results expressed as the number of Disintegrations per Minute (DPM). A Stimulation Index (SI) was calulated for each group. The mean DPM/animals values for the experimental groups treated with the test substance concentrations of 10, 25 and 50% were 693, 1508 and 3275 DPM respectively. The mean DPM value for the control group was 662DPM. The Stimulation Index values calculated for the substance concentrations 10, 25 and 50% were 1, 2.3 and 4.9 respectively. Slight irritation of the ears was shown by all experimental animals at all three concentrations. However, this was not considered to have a toxicologically significant effect on the activity of the nodes. No oedema was observed in any of the animals examined. All auricular lymph nodes of the animals at 50% concentration were considered increased in size. All other auricular lymph nodes were considered normal in size. No macroscopic abnormalities of the surrounding area were noted in any of the animals. Body weights and body weight gain of experimental animals remained in the same range as the controls over the study period. The slight body weight loss in some animals was not considered toxicologically significant. No mortality occurred and no symptoms of systemic toxicity were observed in the animals of the main study. Under the conditions of this study, the test substance, Thioanisole, should be considered to be a skin sensitiser. Based on these results, the test substance should be classified as a Category 1 skin sensitizer and have the hazard statement H317: May casue an allergic reaction associated with it, in accordance with Regulation EC No. 1272/2008. According to Directive 67/548/EEC, the test substance should be classified with the risk phrase R43: May cause sensitization by skin contact and have the symbol Xi, Irritant associated with it.