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Description of key information

Tert-Butylperoxy-3,5,5-trimethylhexanoat was assessed for skin sensitization in a Guinea pig maximization test according to EU method B.6 and OECD guideline 406. Based on the results obtained the test substance was considered to cause skin sensitisation. There was no indication of respiratory sensitisation from occupational use.

Key value for chemical safety assessment

Skin sensitisation

Link to relevant study records
Reference
Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
experimental study
Adequacy of study:
key study
Study period:
1998-06-25 to 1998-07-20
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Justification for type of information:
The study was conducted in 1998 when the in vitro test battery was yet not legally accepted to fullfil the information requirements under REACH.
Qualifier:
according to guideline
Guideline:
OECD Guideline 406 (Skin Sensitisation)
Version / remarks:
1992
Deviations:
no
Qualifier:
according to guideline
Guideline:
EU Method B.6 (Skin Sensitisation)
Version / remarks:
1992
Deviations:
no
GLP compliance:
yes
Type of study:
guinea pig maximisation test
Justification for non-LLNA method:
The study was conducted in 1998 when the GPMT was an accepted and recommended test for assessment of skin sensitization.
Species:
guinea pig
Strain:
Dunkin-Hartley
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River France, 76410 Saint-Aubin-les-Elbeuf, France
- Age at study initiation: 3 months
- Weight at study initiation: males: 364 ± 13 g; females: 360 ± 15 g
- Housing: individually in polycarbonate cages (48 cm x 27 cm x 20 cm) equipped with a polypropylene bottle
- Diet: 106 pellet diet (UAR, 91360 Villemoisson-sur-Orge, France), ad libitum
- Water: drinking water filtered by FG Milipore membrane, ad libitum
- Acclimation period: at least 5 days before the beginning of the study

ENVIRONMENTAL CONDITIONS
- Temperature: 21°C ± 2°C
- Humidity: 30 to 70%
- Air changes (per hr): approximately 12 cycles/ hour of filtered, non-recycled air
- Photoperiod (hrs dark / hrs light): 12 h/ 12 h
Route:
intradermal and epicutaneous
Vehicle:
corn oil
Concentration / amount:
Induction (treated group):
-intradermal injections: LUPEROX 270 at the concentration of 10% (w/w) in corn oil,
- topical application: 0.5 mL LUPEROX 270 undiluted
Route:
epicutaneous, occlusive
Vehicle:
corn oil
Concentration / amount:
Challenge (all groups):
- topical application:0.5 mL LUPEROX 270 undiluted
No. of animals per dose:
treated group: ten males and ten females
control group: five males and five females
Details on study design:
RANGE FINDING TESTS:
By intradermal route:
- 24 hours before treatment, the dorsal region of the animals was clipped,
- intradermal administration of the test substance formulation (0.1 mL) at different concentrations were performed in the interscapular region,
- cutaneous reaction were evaluated approximately 24 and 48 hours and 6 days after the injections.

By cutaneous route:
- 24 hours before treatment, both flank regions of the animals were clipped,
0.5 mL of the undiluted test substance or test substance formulation at the chosen concentration were placed on a dry gauze pad (approximately 4 cm2) which was then applied to the skin and held in place by an occlusive dressing for 24 hours,
- cutaneous reactions were evaluated approximately 24 and 48 hours after removal of the dressings.

MAIN STUDY
A. INDUCTION EXPOSURE
Intradermal route:
On day 1, six injections were made deep into the dermis of a 4 cm x 2 cm clipped interscapular area, using a needle (diameter: 0.5 x 16 mm) mounted on a 1 mL plastic syringe (0.01 mL graduations).

Cutaneous route:
On day 7, the intrascapular area was clipped.
As the test substance was shown to be non-irritant during the preliminary test, the animals were treated with 0.5 mL of sodium lauryl sulphate (10%, w/w) in vaseline on order to induce local irritation.
On day 8, a cutaneous application to the region of the intradermal injections (4 cm x 2 cm) was performed as follows:

Control group: application of 0.5 mL of the vehicle.
Treated group: application of 0.5 mL of the undiluted test substance.

The test substance or the vehicle was placed on a dry gauze pad, which was then applied to the interscapular region.
The pad was held in place for 48 hours by means of an adhesive hypoallergenic dressing and an adhesive an allergenic waterproof plaster.
On removal of the dressing, no residual test substance was observed.
Cutaneous reactions were recorded 1 hour after removal of the occlusive dressing.

B. CHALLENGE EXPOSURE
On day 22, the animals of both groups received an application of 0.5 mL of the undiluted test substance to the posterior right flank and 0.5 mL of the vehicle to the posterior left flank. This application was performed using a 1 mL plastic syringe (0.01 mL) graduations). The test substance or the vehicle was placed on a dry gauze pad, which was then applied to 4 cm2 (2 cm x 2 cm) clipped area of the skin.

The pads were held in contact with the skin for 24 hours by means of an occlusive, hypoallergenic dressing and an adhesive an allergenic waterproof plaster.
On removal of the dressing, no residual test substance was observed.
Challenge controls:
NA
Positive control substance(s):
yes
Remarks:
2,4-Dinitro Chlorobenzene (DNCB) and Mercaptobenzothiazole
Positive control results:
Under this experimental conditions and according to the Magnusson and Kligman method, the test substance DNCB at the concentration of 1 % (w/w) induced positive skin sensitization reactions in 90 % of the guinea-pigs.
Under this experimental conditions and according to the Magnusson and Kligman method, the test substance MERCAPTOBENZOTHIAZOLE at the concentration of 20 % (w/w) induced positive skin sensitization reaction in 30 % of the guinea-pigs.
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
test chemical
Dose level:
10%
No. with + reactions:
18
Total no. in group:
19
Clinical observations:
very slight /well-defined erythema
Key result
Reading:
2nd reading
Hours after challenge:
48
Group:
test chemical
Dose level:
10%
No. with + reactions:
0
Total no. in group:
19
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
negative control
Dose level:
50%
No. with + reactions:
0
Total no. in group:
9
Clinical observations:
none
Key result
Reading:
2nd reading
Hours after challenge:
48
Group:
negative control
Dose level:
50%
No. with + reactions:
0
Total no. in group:
9
Clinical observations:
none
Key result
Group:
positive control
Remarks on result:
not measured/tested
Interpretation of results:
Category 1 (skin sensitising) based on GHS criteria
Conclusions:
Under this experimental conditions and according to the maximization method of Magnusson and Kligman, the test substance tert-Butylperoxy-3,5,5-trimethylhexanoat induces delayed contact hypersensitivity in 18/19 (95%) guinea-pigs.
Executive summary:

Tert-Butylperoxy-3,5,5-trimethylhexanoat was tested to thirty guinea-pig according to the maximization method of Magnusson and Kligman, EU-method B.6 and OECD 406.

Thirty guinea-pigs were allocated to two groups: a control group 1 (five males and five females) and a treated group 2 (ten males and ten females).

On day 1, intradermal injections of Freund's complete adjuvant mixed with the test substance (treated group) or vehicle (control group) were performed in the interscapular region.

On day 7, the same region received a topical application of sodium lauryl sulphate in vaseline (10%, w/w) in order to induce local irritation.

On day 8, the test substance (treated group) or the vehicle (control group) was applied to the same test site which was then covered by an occlusive dressing for 48 hours.

On day 22, after a rest period of 12 days, all animals of the treated and control groups were challenged by a cutaneous application of the test substance (undiluted) to the right flank. The left flank served as control and received the vehicle only. Test substance and vehicle were maintained under an occlusive dressing for 24 hours.

Skin reactions were evaluated approximately 24 and 48 hours after removal of the dressing.

Test substance concentrations were as follows:

Induction (treated group):

-intradermal injections: tert-Butylperoxy-3,5,5-trimethylhexanoat at the concentration of 10% (w/w) in corn oil,

- topical application: 0.5 mL tert-Butylperoxy-3,5,5-trimethylhexanoat undiluted

Challenge (all groups):

- topical application:0.5 mL tert-Butylperoxy-3,5,5-trimethylhexanoat undiluted

At the end of the study, animals were killed without examination of internal organs.

No skin samples were taken from the challenge application sites.

No clinical signs and no deaths related to treatment were noted during the study.

For the challenge application, no cutaneous reactions were observed in the animals of the control group.

In the treated group, at the 24 -hour reading, a very slight or well-defined erythema was noted in 10/10 male and 8/9 female animals.

No cutaneous reactions persisted at the 48 -hour reading.

Dryness of the skin was observed in 2/19 animals at the 24 -hour reading and in all animals at the 48 -hour reading.

The species and strain which were used showed a satisfactory sensitization response in 90% animals treated with DNCB and in 30 % animals treated with Mercaptobenzothiazole.

Under this experimental conditions and according to the maximization method of Magnusson and Kligman, the test substance tert-Butylperoxy-3,5,5-trimethylhexanoat induces delayed contact hypersensitivity in 18/19 (95%) guinea-pigs.

Endpoint conclusion
Endpoint conclusion:
adverse effect observed (sensitising)
Additional information:

Skin sensitisation


The test substance was tested for skin sensitization properties in a Guinea pig maximization test according to EU Method B.6 and OECD guideline 406. No clinical signs and no mortality related to treatment were noted during the study. Following the challenge application, no cutaneous reactions were observed in animals of the control group. In the treated group, at the 24 -hour reading, a very slight and well-defined erythema was noted in 10/19 and 8/19 animals (42 %), respectively. No cutaneous reactions persisted at the 48 -hour reading. Dryness of the skin was observed in 2/19 animals at the 24 -hour reading and in all animals at the 48 -hour reading. According to the maximization method of Magnusson and Kligman, the test substance induces delayed contact hypersensitivity in guinea-pigs.

Respiratory sensitisation

Endpoint conclusion
Endpoint conclusion:
no study available
Additional information:

There was no indication of respiratory sensitisation from occupational use.

Justification for classification or non-classification

Classification, Labelling, and Packaging Regulation (EC) No 1272/2008
The available experimental test data are reliable and suitable for classification purposes under Regulation (EC) No 1272/2008. Based on available data the test item is classified and labelled as skin sensitiser cat 1 (H317 "may cause an allergic skin reaction") according to Regulation (EC) No 1272/2008 (CLP), as amended for the seventeenth time in Regulation (EU) 2021/849.