methylethylketone peroxide trimer

Endpoint summary

Description of key information

It was concluded from 2 independent studies that the overall sensitisation rate was 35%, therefore requiring classification. However, based on the fact that challenge reactions were only observed when using the undiluted test substance (which is irritating), that these reactions were moderate and also seen in controls, and not observed at a concentration of 75%, it can also be concluded that the test substance is a weak sensitiser, if at all.

In study 106/031, at the challenge with undiluted (100%) test substance in both the test and the control animals effects of skin irritation were observed. Since these effects were observed in both test (8/10) and control (3/5) animals and no effects whatsoever were observed in both test and control animals at the challenge concentration of 75% it was concluded that owing to the irritancy of the 100% test material no sensitisation responses could be determined in the 100% group. In the 75% challenge concentration no signs of irritation were observed. The test material was therefore concluded not to cause sensitisation.

However In a second report (106/050) it was concluded that in the first study (106/031) 2/10 test animals showed a more severe reponse than the other animals and controls did. One of these two was killed for humane reasons 24 h after challenge (so that no data were available at 48 h). As 5/10 (50%) of the animals showed slight skin reactions in the second study (106/050) at a challenge of 100% both at 24 h and 48 h after challenge, whereas controls did not, it was concluded that the overall sensitisation rate from these two studies was 7/20 (35%).

In study 189359the skin reactions observed in response to a 50% test substance concentration in five (of the nine) experimental animals in the challenge phase were considered indicative of sensitisation, based on the absence of any response in the control animals. These results indicate a sensitisation rate of 55 per cent.

Therefore, based on a weight of evidence, the substance is considered a skin sensitizer.

Key value for chemical safety assessment

Skin sensitisation

Link to relevant study records

Reference

Reference 1

Endpoint:

skin sensitisation: in vivo (non-LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

August-September 2000

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Well conducted study according to GLP

Reason / purpose for cross-reference:

reference to same study

Qualifier:

according to guideline

Guideline:

OECD Guideline 406 (Skin Sensitisation)

Qualifier:

according to guideline

Guideline:

EU Method B.6 (Skin Sensitisation)

GLP compliance:

yes (incl. QA statement)

Type of study:

guinea pig maximisation test

Justification for non-LLNA method:

LLNA not adopted until 2002

Species:

guinea pig

Strain:

Dunkin-Hartley

Sex:

male

Route:

intradermal and epicutaneous

Vehicle:

arachis oil

Concentration / amount:

Concentration of test material and vehicle used at induction:
- Intradermal induction: 10% w/v in arachis oil BP - Topical induction: undiluted as supplied Concentration of test material and vehicle used for each challenge: Undiluted as supplied and 75% v/v in arachis oil BP

Route:

epicutaneous, occlusive

Vehicle:

arachis oil

Concentration / amount:

Concentration of test material and vehicle used at induction:
- Intradermal induction: 10% w/v in arachis oil BP - Topical induction: undiluted as supplied Concentration of test material and vehicle used for each challenge: Undiluted as supplied and 75% v/v in arachis oil BP

No. of animals per dose:

First study (106/031):
Number of animals in test group: 10
Number of animals in negative control group: 5
Second study (106/050):
Number of animals in test group: 10
Number of animals in negative control group: 5

Challenge controls:

yes

Positive control substance(s):

yes

Remarks:

mercaptobenzothiazole (149-30-4)

Positive control results:

OK

Reading:

1st reading

Hours after challenge:

24

Group:

test chemical

Dose level:

75 %

No. with + reactions:

0

Total no. in group:

10

Remarks on result:

other: Reading: 1st reading. . Hours after challenge: 24.0. Group: test group. Dose level: 75 %. No with. + reactions: 0.0. Total no. in groups: 10.0.

Reading:

1st reading

Hours after challenge:

24

Group:

test chemical

Dose level:

100 %

No. with + reactions:

7

Total no. in group:

10

Remarks on result:

other: Reading: 1st reading. . Hours after challenge: 24.0. Group: test group. Dose level: 100 %. No with. + reactions: 7.0. Total no. in groups: 10.0.

Reading:

2nd reading

Hours after challenge:

48

Group:

test chemical

Dose level:

75 %

No. with + reactions:

0

Total no. in group:

10

Remarks on result:

other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: test group. Dose level: 75 %. No with. + reactions: 0.0. Total no. in groups: 10.0.

Reading:

2nd reading

Hours after challenge:

48

Group:

test chemical

Dose level:

100 %

No. with + reactions:

5

Total no. in group:

10

Remarks on result:

other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: test group. Dose level: 100 %. No with. + reactions: 5.0. Total no. in groups: 10.0.

Reading:

1st reading

Hours after challenge:

24

Group:

negative control

Dose level:

75 %

No. with + reactions:

0

Total no. in group:

5

Remarks on result:

other: Reading: 1st reading. . Hours after challenge: 24.0. Group: negative control. Dose level: 75 %. No with. + reactions: 0.0. Total no. in groups: 5.0.

Reading:

1st reading

Hours after challenge:

24

Group:

negative control

Dose level:

100 %

No. with + reactions:

0

Total no. in group:

5

Remarks on result:

other: Reading: 1st reading. . Hours after challenge: 24.0. Group: negative control. Dose level: 100 %. No with. + reactions: 0.0. Total no. in groups: 5.0.

Reading:

2nd reading

Hours after challenge:

48

Group:

negative control

Dose level:

75 %

No. with + reactions:

0

Total no. in group:

5

Remarks on result:

other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: negative control. Dose level: 75 %. No with. + reactions: 0.0. Total no. in groups: 5.0.

Reading:

2nd reading

Hours after challenge:

48

Group:

negative control

Dose level:

100 %

No. with + reactions:

0

Total no. in group:

5

Remarks on result:

other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: negative control. Dose level: 100 %. No with. + reactions: 0.0. Total no. in groups: 5.0.

In the first study (106/031), at the challenge with undiluted (100%) test substance in both the test and the control animals effects of skin irritation were observed. Since these effects were observed in both test (8/10) and control (3/5) animals and
no effects whatsoever were observed in both test and control animals at the challenge concentration of 75% it was
concluded that owing to the irritancy of the 100% test material no sensitisation responses could be determined in
the 100% group. In the 75% challenge concentration no signs of irritation were observed. The test material was therefore
concluded not to cause sensitisation.

In the second report (106/050) it was, however, concluded that in the first study (106/031) 2/10 test animals showed a more severe reponse than the other animals and controls did. One of these two was killed for humane reasons 24 h after challenge (so that no data were available at 48 h).

As 5/10 (50%) of the animals showed slight skin reactions in the second study (106/050) at a challenge of 100% both at 24 h and 48 h after challenge, whereas controls did not, it was concluded that the overall sensitisation rate from these two studies was 7/20 (35%).

Signs of irritation during induction:
- Intradermal induction: Well defined to moderate to  severe erythema observed in all test group animals.
- Topical induction: Very slight to well-defined erythema with very slight oedema was noted in all test group animals at the 1 hour observation. In twelve test group animals very slight  to well-defined erythema was noted at the induction
sites after 24 hours.

Interpretation of results:

sensitising

Conclusions:

Under the conditions of the test, the test material produced a 50% (5/10) sensitisation rate. The results of this study can be collated with the results of the previous study (SPL Project Number 106/031) to give a total sensitization rate of 35% (7/20)

Executive summary:

Under the conditions of the test, the test material produced a 50% (5/10) sensitisation rate.  The results of this study can be collated with the results of the previous study (SPL Project Number 106/031) to give a total sensitization rate of 35% (7/20)

Reference 2

Endpoint:

skin sensitisation: in vivo (non-LLNA)

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Apparently well conducted GLP study. No certificate of analysis.

Qualifier:

according to guideline

Guideline:

OECD Guideline 406 (Skin Sensitisation)

GLP compliance:

yes (incl. QA statement)

Type of study:

guinea pig maximisation test

Justification for non-LLNA method:

LLNA not adopted until 2002

Species:

guinea pig

Strain:

Himalayan

Sex:

female

Details on test animals and environmental conditions:

Air-conditioned room with approximately 15 air changes per hour and the
environment controlled with optimal conditions considered as being a temperature
of 21°C and a relative humidity of 50%. Fluctuations from these optimal
conditions were noted, but were considered not to have affected study integrity.
Lighting was 12 hours artificial fluorescent light and 12 hours dark per day.

Accommodation:
Group housing of 5 animals per labelled metal cage with wire-mesh floors and
equipped with an automatic drinking system (ITL, Bergen, The Netherlands). The
acclimatisation period was at least 5 days before the start of treatment under
laboratory conditions.

Diet:
Free access to standard guinea pig diet, including ascorbic acid (1600 mg/kg);
LC 23-B, pellet diameter 4mm (Hope farms, Woerden, The Netherlands).
Certificates of analysis were examined and retained in the NOTOX archives.
In addition, hay (B.M.I., Helmond, The Netherlands) was provided once a week.

Water:
Free access to tap-water, diluted with decalcified water. Certificates of
quarterly analysis for tap-water were examined and retained in the NOTOX
archives.

Route:

intradermal and epicutaneous

Vehicle:

corn oil

Concentration / amount:

Based on the results, the test substance concentrations selected for the Main
Study were a 20% concentration for the intradermal induction and the undiluted
test substance for the epidermal induction exposure.
A 50% and 20% test substance concentration were selected for the challenge
phase.

Route:

epicutaneous, semiocclusive

Vehicle:

corn oil

Concentration / amount:

Based on the results, the test substance concentrations selected for the Main
Study were a 20% concentration for the intradermal induction and the undiluted
test substance for the epidermal induction exposure.
A 50% and 20% test substance concentration were selected for the challenge
phase.

No. of animals per dose:

5 control and 10 exposed

Details on study design:

INDUCTION: Experimental animals

Day 1 The scapular region was clipped and three pairs of intradermal injections
(0.1 ml/site) were made in this area as follows:
A) Freunds' Complete Adjuvant (Difco, Detroit, U.S.A.), 50:50 with water
for injection (Fresenius AG, Bad Homburg, Germany).
B) The test substance at a 20% concentration.
C) A 1:1 mixture of the test substance, at twice the concentration used
in (B), and Freunds' Complete Adjuvant.
Note: One of each pair was on each side of the midline and from cranial A) to caudal C).
Day 3 The dermal reactions caused by the intradermal injections were assessed
for irritation.
Day 8 The scapular area between the injection sites was clipped and
subsequently treated with 0.5 ml of the undiluted test substance using a
Scotchpak-non-woven patch (2x3 cm) mounted on Micropore tape and secured
with Coban elastic bandage.
The dressing was removed after 48 hours exposure, the skin cleaned of
residual test substance and the dermal reactions caused by the epidermal
exposure were assessed for irritation.

INDUCTION - CONTROL ANIMALS
The control animals were treated as described for the experimental
animals, except that, instead of the test substance, the vehicle was
administered.

CHALLENGE: All animals
Day 22 One flank of all animals was clipped and treated by epidermal application
of a 50% and 20% test SUbstance concentration and the vehicle (0.5 ml
each), using Scotchpak-non-woven patches (2x3 cm) mounted on Micropore
tape and secured with Cob an elastic bandage.
The dressing was removed after 24 hours exposure and the skin cleaned of
residual test substance and vehicle. The treated sites were assessed for
challenge reactions 24 and 48 hours after removal of the dressing.

Positive control substance(s):

yes

Remarks:

ALPHA-HEXYLCINNAMICALOEHYDE, TECH. 85%

Positive control results:

The skin reactions in the experimental animals observed in response to the 50%
test substance concentration in the challenge phase were considered indicative
of sensitisation, based on the intensity of the responses compared to those seen
in the control animals.
These results lead to a sensitisation rate of 100 percent.
From these results, it was concluded that the females of the albino Himalayan
strain of guinea pig is an appropriate animal model for the performance of
studies designed to evaluate the sensitising potential of a substance in a
Maximisation type of test.

Reading:

1st reading

Hours after challenge:

48

Group:

test chemical

Dose level:

50%

No. with + reactions:

5

Total no. in group:

10

Clinical observations:

None

Remarks on result:

other: Reading: 1st reading. . Hours after challenge: 48.0. Group: test group. Dose level: 50%. No with. + reactions: 5.0. Total no. in groups: 10.0. Clinical observations: None.

Interpretation of results:

sensitising

Remarks:

Criteria used for interpretation of results: EU

Conclusions:

The skin reactions observed in response to a 50% test substance concentration in
five (of the nine) experimental animals in the challenge phase were considered
indicative of sensitisation, based on the absence of any response in the control
animals.
These results indicate a sensitisation rate of 55 per cent.

Executive summary:

Assessment for Contact Hypersensitivity to 0-129 in the Albino Guinea Pig (Maximization Test). The study was carried out in accordance with the DECO Guideline No. 406, 'Skin Sensitisation', the EEC Directive 92f69fEEC, Part B.6, 'Skin Sensitisation' and based on the method described by Magnusson and Kligman, 'Allergic Contact Dermatitis in the Guinea Pig - Identification of Contact Allergens'. Test substance concentrations (in corn oil) selected for the Main study were based on the results of a preliminary study. In the Main study, ten experimental animals were intradermally injected with a 20% concentration and epidermally exposed to the undiluted test substance. Five control animals were similarly treated, but with vehicle only. Two weeks after the epidermal application all animals were challenged with a 50% and 20% test substance concentration and the vehicle. Skin reactions varying between grades 1 and 2, were observed in five experimental .animals in response to the 50% test substance concentration 48 hours after the challenge exposure. No skin reactions were evident to.the 20% test substance concentration in the experimental animals. No skin reactions were evident in the control animals (see Table 3). The skin reactions observed in response to a 50% test substance concentration in five (of the nine) experimental animals in the challenge phase were considered indicative of sensitisation, based on the absence of any response in the control animals. These results indicate a sensitisation rate of 55 per cent.

Reference 3

Endpoint:

skin sensitisation: in vivo (non-LLNA)

Type of information:

experimental study

Adequacy of study:

weight of evidence

Study period:

1997

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: GLP study. Undiluted test article was used for dermal induction and challenge which was an irritating concentration.

Qualifier:

according to guideline

Guideline:

OECD Guideline 406 (Skin Sensitisation)

GLP compliance:

yes (incl. QA statement)

Type of study:

guinea pig maximisation test

Justification for non-LLNA method:

LLNA not adopted until 2002

Species:

guinea pig

Strain:

Dunkin-Hartley

Sex:

male

Details on test animals and environmental conditions:

Twenty-four male albino Dunkin Hartley guinea pigs supplied by David Hall
Limited, Burton-on-Trent, Staffordshire, UK were used. At the start of the main
study the animals weighed 341 to 419g, and were approximately eight to
twelve weeks old. After an acclimatisation period of at least five days, each
animal was selected at random and given a number unique within the study
which was written on a small area of clipped rump using a black indelible
marker-pen.

The animals were housed singly or in pairs in solid-floor polypropylene cages
furnished with wood flakes. Free access to mains tap water and food (Guinea
Pig FD1 Diet, Special Diets Services Limited, Witham, Essex, UK) was allowed
throughout the study.

The animal room was maintained at a temperature of 19 to 22· C and relative
humidity of 34 to 60%. The rate of air exchange was approximately fifteen
changes per hour and the lighting was controlled by a time switch to give
twelve hours continuous light and twelve hours darkness.

Route:

intradermal and epicutaneous

Vehicle:

arachis oil

Concentration / amount:

Intradermal Induction: 10% w/v in arachis oil BP
10% w/v in a mixture of Freund's
Complete Adjuvant plus distilled water

Topical Induction : undiluted as supplied

Topical Challenge: undiluted as supplied and 75% v/v in arachis oil BP

Route:

epicutaneous, occlusive

Vehicle:

arachis oil

Concentration / amount:

Intradermal Induction: 10% w/v in arachis oil BP
10% w/v in a mixture of Freund's
Complete Adjuvant plus distilled water

Topical Induction : undiluted as supplied

Topical Challenge: undiluted as supplied and 75% v/v in arachis oil BP

No. of animals per dose:

10

Details on study design:

Induction of the 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:
a) Freund's Complete Adjuvant plus distilled water in the ratio 1: 1
b) a 10% w/v solution of the test material in arachis oil BP
c) a 10% w/v emulsion of the test material in a 1:1 preparation of
Freund's Complete Adjuvant plus distilled water.

Approximately 24 and 48 hours after intradermal injection the
degree of erythema at the test material injection sites (ie. injection
site b) was evaluated according to the scale shown in Appendix X.
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. A filter
paper patch (WHATMAN No.4: approximate size 40 mm x 20 mm),
saturated with the undiluted test material was applied to the
prepared skin and held in place with a strip of surgical adhesive
tape (BLENDERM: approximate size 50 mm x 30 mm) covered with
an overlapping length of aluminium foil. The patch and foil were
further secured with 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.
The degree of erythema and oedema was quantified one and twentyfour
hours following removal of the patches.
Any other reactions were also recorded.

Induction of the Control Animals: Intradermal injections were
administered using an identical procedure to that used for the test
animals, except that the injections were:
a) Freund's Complete Adjuvant plus distilled water in the ratio 1:1
b) arachis oil BP
c) a 50% w/v formulation of arachis oil BP in Freund's Complete
Adjuvant/distilled water 1:1
Approximately 24 and 48 hours after intradermal injection the
degree of erythema at the vehicle injection sites (ie injection site b)
was evaluated.

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.

Challenge
Shortly before treatment on Day 21, an area of approximately
50 mm x 70 mm on both flanks of each animal, was clipped free of
hair with veterinary clippers.
A square filter paper patch (WHATMAN No.4: approximate size
20 mm x 20 mm), saturated with the undiluted test material was
applied to the shorn right flank of each animal and was held in
place with a strip of surgical adhesive tape (BLENDERM:
approximate size 40 mm x 50 mm). To ensure that the maximum
non-irritant concentration was used at challenge, the test material at
a concentration of 75% v/v in arachis oil BP was similarly applied
to a skin site on the left shorn flank. The patches were occluded
with an overlapping length of aluminium foil and secured with 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 challenge sites were swabbed
with cotton wool soaked in diethyl ether to remove residual
material. The position of the treatment sites was identified by using
a black indelible marker-pen. Due to the regrowth of fur the
challenge sites of test and control group animals were depilated
prior to the 24-hour observation.

Challenge controls:

See above.

Positive control substance(s):

yes

Remarks:

2·Mercaptobenzothiazole

Positive control results:

90% positive

Reading:

1st reading

Hours after challenge:

24

Group:

test chemical

Dose level:

75%

No. with + reactions:

0

Total no. in group:

10

Clinical observations:

None

Remarks on result:

other: Reading: 1st reading. . Hours after challenge: 24.0. Group: test group. Dose level: 75%. No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: None.

Interpretation of results:

sensitising

Remarks:

Criteria used for interpretation of results: EU

Conclusions:

In this study (106/031), at the challenge with undiluted (100%) test substance in both the test and the control animals effects of skin irritation were observed. Since these effects were observed in both test (8/10) and control (3/5) animals and no effects whatsoever were observed in both test and control animals at the challenge concentration of 75% it was concluded that owing to the irritancy of the 100% test material no sensitisation responses could be determined in the 100% group. In the 75% challenge concentration no signs of irritation were observed. The test material was therefore concluded not to cause sensitisation.

However In a second report (106/050) it was concluded that in the first study (106/031) 2/10 test animals showed a more severe reponse than the other animals and controls did. One of these two was killed for humane reasons 24 h after challenge (so that no data were available at 48 h).
As 5/10 (50%) of the animals showed slight skin reactions in the second study (106/050) at a challenge of 100% both at 24 h and 48 h after challenge, whereas controls did not, it was concluded that the overall sensitisation rate from these two studies was 7/20 (35%).

Executive summary:

A study was performed to assess the contact sensitisation potential of the test material in the albino guinea pig. The study was performed in compliance with the OECD Guidelines for Testing of Chemicals No. 406 "Skin Sensitisation" (adopted 17 July 1992) and Method B6 of Commission Directive 92/69/EEC (which constitutes Annex V of Council Directive 67/548/EEC).

Ten test and five 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-10% w/v in arachis oil BP;

Topical Induction- undiluted as supplied Topical Challenge- undiluted as supplied and 75% v/v in arachis oil BP In this study (106/031), at the challenge with undiluted (100%) test substance in both the test and the control animals effects of skin irritation were observed. Since these effects were observed in both test (8/10) and control (3/5) animals and no effects whatsoever were observed in both test and control animals at the challenge concentration of 75% it was concluded that owing to the irritancy of the 100% test material no sensitisation responses could be determined in the 100% group. In the 75% challenge concentration no signs of irritation were observed. The test material was therefore concluded not to cause sensitisation. However In a second report (106/050) it was concluded that in the first study (106/031) 2/10 test animals showed a more severe reponse than the other animals and controls did. One of these two was killed for humane reasons 24 h after challenge (so that no data were available at 48 h). As 5/10 (50%) of the animals showed slight skin reactions in the second study (106/050) at a challenge of 100% both at 24 h and 48 h after challenge, whereas controls did not, it was concluded that the overall sensitisation rate from these two studies was 7/20 (35%).

Reference 4

Endpoint:

skin sensitisation: in vitro

Data waiving:

study scientifically not necessary / other information available

Justification for data waiving:

other:

Endpoint conclusion

Endpoint conclusion:

adverse effect observed (sensitising)

Justification for classification or non-classification

Based on a weight of evidence from three studies, the substance meets the critieria for classificatin as a skin sensitizer.