Hexadecyl 3,5-bis-tert-butyl-4-hydroxybenzoate

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

13 - 25 June 2012

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: This study has been performed according to OECD and/or EC guidelines and according to GLP principles.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2012

Report date:

2012

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.42 (Skin Sensitisation: Local Lymph Node Assay)

Deviations:

no

Test guideline 3

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.2600 (Skin Sensitisation)

Deviations:

no

GLP compliance:

yes

Type of study:

mouse local lymph node assay (LLNA)

Test material

Test material information

Constituent 1

Reference substance name:

CYASORB® UV-2908 Light Stabilizer

IUPAC Name:

CYASORB® UV-2908 Light Stabilizer

Details on test material:

- Name of test material (as cited in study report):CYASORB® UV-2908 Light Stabilizer
- Substance type: White to off-white powder
- Physical state: powder
- Purity: >98%
- Lot/batch No.: WP1022803
- Expiration date of the lot/batch: 18 April 2013
- Storage condition of test material: At room temperature protected from light

In vivo test system

Test animals

Species:

mouse

Strain:

other: CBA/J

Sex:

female

Details on test animals and environmental conditions:

- Source: Janvier, Le Genest-Saint-Isle, France
- Age at study initiation: Young adult animals (approx. 9-10 weeks old)
- Weight at study initiation: Body weight variation was within +/- 20% of the sex mean (19-23 grams)
- Housing: Animals were group housed in labeld makrolon cages.
- Diet: Free access to pelleted rodent diet (SM R/M-Z from SSNIFF® Spezialdiäten GmbH, Soest, Germany).
- Water: Free access to tap water.
- Acclimation period: At least 5 days

ENVIRONMENTAL CONDITIONS
Environmental controls for the animal room were set to maintain 18 to 24°C, a relative humidity of 40 to 70%, approximately 15 room air changes/hour, and a 12-hour light/12-hour dark cycle. Any variations to these conditions were maintained in the raw data and had no effect on the outcome of the study.

IN-LIFE DATES: From13 to 25 June 2012

Study design: in vivo (LLNA)

Vehicle:

methyl ethyl ketone

Concentration:

0, 10, 25 , 50%

No. of animals per dose:

5

Details on study design:

The vehicle was selected based on trial formulations performed at NOTOX and on test substance data supplied by the sponsor.

RANGE FINDING TESTS:
In the interest of animal welfare and to minimize any testing likely to produce severe responses in animals, a weight of evidence analysis was performed prior to start of this study. All available information was evaluated (e.g. existing human and animal data, literature, substance data supplied by the sponsor, analysis of structure activity relationships (SAR), physicochemical properties and reactivity (pH, buffering capacity).

MAIN STUDY
ANIMAL ASSIGNMENT AND TREATMENT
- Name of test method: Local Lymph Node Assay
- Criteria used to consider a positive response: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 substance may be regarded as a skin sensitizer. The results were evaluated according to the Globally Harmonized System of Classification and Labelling of Chemicals (GHS) of the United Nations (20011) and the Regulation (EC) No 1272/2008 of the European Parliament and of the Council of 16 December 2008 on Classification, Labelling and Packaging of substances and mixtures.

ANIMAL ASSIGNMENT
Three groups of five animals were treated with one test substance concentration per group. One group of five animals was treated with vehicle.

TREATMENT PREPARATION AND ADMINISTRATION:
Test substance preparation: The test substance formulations (w/w) were prepared within 4 hours prior to each treatment. Homogeneity was obtained to visually acceptable levels.
Rationale for vehicle: The vehicle was selected based on trial formulations performed at NOTOX and on test substance data supplied by the sponsor.

Induction - Days 1, 2 and 3; Excision of nodes - Day 6; Tissue processing for radioacitivity - Day 6; Radioactivity measurements - Day 7; Performed according to test guidelines.

Observations:
Mortality/Viability: Twice daily.
Body weights: On Day 1 (pre-dose) and Day 6 (prior to necropsy).
Clinical signs: Once daily on Days 1-6 (on Days 1-3 between 3 and 4 hours after dosing).
Irritation: Once daily on Days 1-6 (on Days 1 - 3 immediately after dosing) according to the following numerical scoring system. Furthermore, a description of all other (local) effects was recorded according to guidelines.

Necropsy: No necropsy was performed according to protocol.

Positive control substance(s):

hexyl cinnamic aldehyde (CAS No 101-86-0)

Statistics:

Not performed.

Results and discussion

Positive control results:

The six-month reliability check with Alpha-hexylcinnamicaldehyde indicates that the Local Lymph Node Assay as performed at NOTOX is an appropriate model for testing for contact hypersensitivity. See attached document 'Reliability check'.

In vivo (LLNA)

Results

Results 1

Parameter:

SI

Value:

1.7

Test group / Remarks:

10% (w/w)

Results 2

Parameter:

SI

Value:

0.9

Test group / Remarks:

25% (w/w)

Results 3

Parameter:

SI

Value:

1.2

Test group / Remarks:

50% (w/w)

Results 4

Parameter:

other: disintegrations per minute (DPM)

Value:

557

Test group / Remarks:

10% (w/w)

Results 5

Parameter:

other: disintegrations per minute (DPM)

Value:

284

Test group / Remarks:

25% (w/w)

Results 6

Parameter:

other: disintegrations per minute (DPM)

Value:

380

Test group / Remarks:

50% (w/w)

Results 7

Parameter:

other: disintegrations per minute (DPM)

Value:

323

Test group / Remarks:

Vehicle control

Any other information on results incl. tables

Results Pre-screen test:

At a 25 and 50% test substance concentration no signs of systemic toxicity were noted and the irritation was maximally grade 2 (only on Day 3). Variations in ear thickness during the observation period were less than 25% from Day 1 pre-dose values. Based on these results, the highest test substance concentration selected for the main study was a 50% concentration.

Other results - main study:

Skin reactions / Irritation: No irritation of the ears was observed in any of the animals examined.

White test substance remnants were noted, which did not hamper scoring for erytema.

Systemic toxicity/Body weights:

No mortality occurred and no clinical signs of systemic toxicity were observed in the animals of the main study. Body weights and body weight gain of experimental animals remained in the same range as controls over the study period. The body weight loss noted for some animals across the dose groups was considered not toxicologically significant since the changes were slight in nature and no concentration-related incidence was apparent.

Macroscopy of the auricular lymph nodes and surrounding area:

All auricular lymph nodes of the animals of the experimental and control groups were considered normal in size. No macroscopic abnormalities of the surrounding area were noted in any of the animals.

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

In an LLNA study with mice, SI values calculated for the substance concentrations 10, 25 and 50% were 1.7, 0.9 and 1.2 respectively

Since there was no indication that the test substance elicits an SI ≥ 3 when tested up to 50%, CYASORB® UV-2908 Light Stabilizer was considered not to be a skin sensitizer. It was established that the EC3 value (the estimated test substance concentration that will give a SI =3) (if any) exceeds 50%.

Based on these results, CYASORB® UV-2908 Light Stabilizer would not be regarded as a skin sensitizer according to the recommendations made in the test guidelines. The test substance does not have to be classified and has no obligatory labelling requirement for sensitization by skin contact according to the Globally Harmonized System of Classification and Labelling of Chemicals (GHS) of the United Nations (2011) and the Regulation (EC) No 1272/2008 on classification, labelling and packaging of substances and mixtures.