Zinc 3,5-bis(α-methylbenzyl)salicylate

Endpoint summary

Key value for chemical safety assessment

Skin sensitisation

Link to relevant study records

Reference

Endpoint:

skin sensitisation: in vivo (non-LLNA)

Type of information:

migrated information: read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Study period:

Study was performed between 24 August 1992 and 3 October 1992

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: OECD guideline and GLP compliant study. Read-across study therefore categorised as Klimisch 2.

Reason / purpose for cross-reference:

reference to same study

Reason / purpose for cross-reference:

reference to other study

Qualifier:

according to guideline

Guideline:

OECD Guideline 406 (Skin Sensitisation)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.6 (Skin Sensitisation)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Type of study:

guinea pig maximisation test

Species:

guinea pig

Strain:

Dunkin-Hartley

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: David Hall Limited, Burton-on-Trent, Staffordshire, U.K.
- Age at study initiation: approximately 8 to 12 weeks old
- Weight at study initiation: 310 - 413g
- Housing: The animals were housed in groups of up to three in solid-floor polypropylene cages furnished with softwood shavings.
- Diet (e.g. ad libitum): Free access food (Guinea Pig FD1 Diet, Special Diet Services Limited, Witham, Essex, U.K.) was allowed throughout the study.
- Water (e.g. ad libitum): Free access to mains tap waterwas allowed throughout the study.
- Acclimation period: at least f five days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19 - 23
- Humidity (%): 51 - 75
- Air changes (per hr): approximately 15
- Photoperiod (hrs dark / hrs light): 12 hours light and 12 hours darkness.

On one occasion the humidity was above the limit specified in the protocol (70%). This was considered not to have affected the purpose or integrity of the study.

Route:

intradermal

Vehicle:

water

Remarks:

Distilled water

Concentration / amount:

Based on the results of sighting tests, the concentrations of test material for the induction and challenge phases were selected as follows:
- Intradermal Induction: 5% (w/v) in distilled water
- Topical Induction: undiluted as supplied
- Topical Challenge: 75% and 50% (v/v) in distilled water

Route:

epicutaneous, occlusive

Vehicle:

water

Remarks:

Distilled water

Concentration / amount:

Based on the results of sighting tests, the concentrations of test material for the induction and challenge phases were selected as follows:
- Intradermal Induction: 5% (w/v) in distilled water
- Topical Induction: undiluted as supplied
- Topical Challenge: 75% and 50% (v/v) in distilled water

No. of animals per dose:

20 animals for test substance
10 animals for control

Details on study design:

RANGE FINDING TESTS:
Selection of Concentrations for Main Study (Sighting Tests): The concentrations of test material to be used at each stage of the main study were determined by ‘sighting tests’ in which groups of guinea pigs were treated with various concentrations of test material. The procedures were as follows:
a) Selection of Concentration for Intradermal Induction: Four animals were intradermally injected with preparations of test material (1%, 5%, 10% or 25% w/v in distilled water). The highest concentration that did not cause local necrosis, ulceration or systemic toxicity, was selected for the intradermal induction stage of the main study.
b) Selection of Concentration for Topical Induction: Two guinea pigs (intradermally injected with Freund’s Complete Adjuvant eight days earlier) were treated with the undiluted test material and three preparations of the test material (75%, 50% and 25% v/v in distilled water). The highest concentration producing only mild to moderate dermal irritation after a 48-hour occlusive exposure, was selected for the topical induction stage of the main study.
c) Selection of Concentration for Topical Challenge: The undiluted test material andthree preparations of the test material (75%, 50% and 25% v/v in distilled water) were applied occlusively to the flanks of two guinea pigs for a period of 24 hours. These guinea pigs did not form part of the main study but had been treated identically to the control animals of the main study, up to Day 14. The highest non-irritant concentration of the test material and one lower concentration were selected for the topical challenge stage of the main study.

MAIN STUDY
A. INDUCTION EXPOSURE:
Test Animals: Shortly before treatment on Day 0 the hair was removed from an area approximately 40 mm x 60 mm on the shoulder region of each animal with veterinary clippers. A row of three injections (0.1 ml each) was made on each side of the mid-line. The injections were:
1) Freund’s Complete Adjuvant plus distilled water in the ratio 1:1.
2) a 5% (w/v) dilution of test material in distilled water.
3) a 5% (w/v) dilution of test material in a 1:1 preparation of Freund’s Complete Adjuvant plus distilled water.

One week later (Day 7), the same area on the shoulder region used previously for intradermal injections was clipped again and treated with a topical application of the undiluted test material. The undiluted test material (0.2 - 0.3 ml) was applied on filter paper (WHATMAN No.4: approximate size 40 mm x 20 mm) which was held in place by a strip of surgical adhesive tape (BLENDERM: approximate size 60 mm x 25mm) and covered with an overlapping length of aluminium foil. The patch and foil were further secured by a strip of elastic adhesive bandage (ELASTOPLAST: approximate size 250 mm x 35 mm) wound in a double layer around the torso of each animal, This occlusive dressing was kept in place for 48 hours.

Erythematous reactions were quantified one and twenty-four hours following removal of the patches using the 0 - 3 scale shown below:
0 - no reaction
1 - scattered mild redness
2 - moderate and diffuse redness
3 - intense redness and swelling

Control Animals: Intradermal injections were administered using an identical procedure to that used for the test animals, except that the injections were:
1) Freund’s Complete Adjuvant plus distilled water in the ratio 1:1.
2) distilled water.
3) Freund’s Complete Adjuvant plus distilled water in the ratio 1:1

The topical applications followed the same procedure as for the test animals except that nothing was applied to the filter paper. Skin reactions were quantified as for the test animals.

B. CHALLENGE EXPOSURE:
Shortly before treatment on Day 21, an area, approximately 50 mm x 70 mm on both flanks of each animal, was clipped free of hair with veterinary clippers.

A quantity of 0.1 - 0.2 ml of the test material formulation (75% v/v in distilled water) was applied to the shorn right flank of each animal on a square of filter paper (WHATMAN No.4: approximate size 20 mm x 20 mm) which was held in place by a strip of surgical adhesive tape (BLENDERM: approximate size 40 mm x 50 mm). To ensure that that maximum non-irritant concentration was used at challenge, the test material at a concentration of 50% (v/v) in distilled water was also similarly applied to a separate skin site on the right shorn flank. The vehicle alone was similarly applied to the left shorn flank. The patches were occluded with an overlapping length of aluminium foil and secured by a strip of elastic adhesive bandage (ELASTOPLAST: approximate size 250 mm x 75 mm) wound in a double layer around the torso of each animal.

After 24 hours, the dressing was carefully cut using blunt- tipped scissors, removed and discarded. The treatment sites were decontaminated with distilled water and the position of the treatment sites identified by using a black indelible marker-pen. On Day 23 the treatment sites were clipped free of hair with veterinary clippers.

OTHER:
Evaluation of Skin Reactions: Approximately 24 and 48 hours after challenge dressing removal erythematous reactions were quantified using the four-point scale below:

0 - no reaction
1 - scattered mild redness
2 - moderate and diffuse redness
3 - intense redness and swelling

Evaluation of data: The percentage of test animals that showed a more severe reaction at the test material challenge site than the most severe reaction seen in the control animals, was compared with the following scale:

% of animals Classification of
sensitised: sensitisation potential:

0 non-sensitiser
> 0 - 8 weak sensitiser
> 8 - 28 mild sensitiser
> 28 - 64 moderate sensitiser
> 64 80 strong sensitiser
> 80 - 100 extreme sensitiser

A response in at least 30% of the animals is considered positive and the risk phrase R 43 “MAY CAUSE SENSITISATION BY SKIN CONTACT” is required.

Positive control substance(s):

yes

Remarks:

2,4 dinitro chlorobezene in arachis oil

Positive control results:

produced an 89% sensitization rate.

Reading:

1st reading

Hours after challenge:

24

Group:

test chemical

Dose level:

75% and 50%

No. with + reactions:

0

Total no. in group:

20

Remarks on result:

other: Reading: 1st reading. . Hours after challenge: 24.0. Group: test group. Dose level: 75% and 50%. No with. + reactions: 0.0. Total no. in groups: 20.0.

Reading:

2nd reading

Hours after challenge:

48

Group:

test chemical

Dose level:

75% and 50%

No. with + reactions:

0

Total no. in group:

20

Remarks on result:

other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: test group. Dose level: 75% and 50%. No with. + reactions: 0.0. Total no. in groups: 20.0.

Reading:

1st reading

Hours after challenge:

24

Group:

negative control

Dose level:

0%

No. with + reactions:

0

Total no. in group:

9

Remarks on result:

other: Reading: 1st reading. . Hours after challenge: 24.0. Group: negative control. Dose level: 0%. No with. + reactions: 0.0. Total no. in groups: 9.0.

Reading:

2nd reading

Hours after challenge:

48

Group:

negative control

Dose level:

0%

No. with + reactions:

0

Total no. in group:

9

Remarks on result:

other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: negative control. Dose level: 0%. No with. + reactions: 0.0. Total no. in groups: 9.0.

Reading:

1st reading

Hours after challenge:

24

Group:

positive control

Dose level:

1% DNCB

No. with + reactions:

16

Total no. in group:

18

Remarks on result:

other: Reading: 1st reading. . Hours after challenge: 24.0. Group: positive control. Dose level: 1% DNCB. No with. + reactions: 16.0. Total no. in groups: 18.0.

a) Skin Reactions Observed After Topical Induction (See Appendices IV and V of attached report for full results):

- One control animal was found dead on day 9. The cause of death was not determined but the absence of this animal was not considered to affect the purpose or integrity of the study.

- Scattered mild redness and moderate and diffuse redness were elicited by the test material. Residual test material was noted at the test sites of all test animals one and 24 hours after dressing removal.

 

b) Skin Reactions Observed After Topical Challenge (See Tables 1 and 2 of attached report for full results):

- No adverse reactions were noted at the test material and vehicle control sites of the test or control animals at the 24 and 48-hour observations.

 

c) See Appendices VI and VII of attached report for individual body weights and body weight gains of test animals.

Interpretation of results:

not sensitising

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

The test material, a mixture containing zinc 3,5-bis(α-methylbenzyl)salicylate, produced a 0% (0/20) sensitisation rate and was classified as a NON-SENSITISER to guinea pig skin.
The test material was also classified as a non-sensitiser according to EEC labelling regulations. No risk phrase is therefore required.

Executive summary:

A study was performed to assess the skin contact sensitisation potential of the test material in the albino guinea pig.

Twenty test and ten control animals were used for the main study. Based on the results of sighting tests, the concentrations of test material for the induction and challenge phases were selected as follows:

 

Intradermal Induction: 5% (w/v) in distilled water

Topical Induction: undiluted as supplied

Topical Challenge: 75% and 50% (v/v) in distilled water

 

The test material produced a 0% (0/20) sensitisation rate and was classified as a non-sensitiser to guinea pig skin. The test material is therefore not classified as a sensitiser according to the CLP Regulation.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not sensitising)

Additional information:

The test material, a mixture containing 30.2% zinc 3,5-bis(alpha-methylbenzyl)salicylate, produced a 0% (0/20) sensitisation rate in guinea pigs therefore, by read-across, zinc 3,5-bis(α-methylbenzyl)salicylate is not classified as a sensitiser in accordance with the CLP Regulation.

Justification for read-across

This study was conducted using a liquid mixture containing 30.2% zinc 3,5-bis(alpha-methylbenzyl)salicylate.

The remaining substances present in the mixture include:

-         5.5% PSMS-11

-         1.1% PVA

-         0.7% DIBLAME

-         < 1% Other

-         Remainder: water

PSMS-11 and PVA are not considered to be hazardous substances. DIBLAME is classified as H315: Causes skin irritation, H317: May cause an allergic skin reaction and H411: Toxic to aquatic life with long lasting effects. However, the concentration of DIBLAME in the mixture is below that which triggers classification of the mixture for these hazards. Therefore, it is considered justified to read-across from this mixture to zinc 3,5-bis(α-methylbenzyl)salicylate.

Migrated from Short description of key information:
By read-across, zinc 3,5-bis(α-methylbenzyl)salicylate, is not classified as a sensitiser in accordance with the CLP Regulation.

Justification for selection of skin sensitisation endpoint:
Only 1 study is available.

Respiratory sensitisation

Endpoint conclusion

Endpoint conclusion:

no study available

Justification for classification or non-classification

The above study has been ranked reliability 2 according to the Klimisch et al system. This ranking was deemed appropriate because it is a read-across, GLP study conducted according to a recognised test method. Sufficient dose ranges and numbers are detailed; hence it is appropriate for use based on reliability and animal welfare grounds.

The above results triggered no classification in accordance with the CLP Regulation (EC No 1272/2008).