mecoprop-P (ISO); (R)-2-(4-chloro-2-methylphenoxy)propionic acid

Endpoint summary

Description of key information

Rees (1995)

Under the conditions of this study, repeated administration of the test material did not cause delayed contact hypersensitivity in the guinea-pig.

Supporting Study: Rossbacher & Hellwig (1995)

Under the conditions of the study the test material does not have a sensitising effect on the skin of the guinea pig in the Maximisation Test.

Skin Sensitisation In Vitro: Waiver

An in vitro skin sensitisation study does not need to be conducted because adequate data from in vivo skin sensitisation studies are available.

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:

11 August 1994 to 18 September 1994

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 406 (Skin Sensitisation)

Version / remarks:

1981

Deviations:

no

Qualifier:

according to guideline

Guideline:

EPA OPP 81-6 (Skin Sensitisation)

Version / remarks:

1984

Deviations:

no

GLP compliance:

yes

Type of study:

guinea pig maximisation test

Justification for non-LLNA method:

Data from existing non-LLNA study available.

Specific details on test material used for the study:

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing: The test material was prepared at appropriate concentrations in propylene glycol or in propylene glycol in Freunds Complete Adjuvant. Concentrations of the test material were expressed gravimetrically, in terms of the test material as received. Fresh doses were prepared on the day of administration and any unused material was disposed of on the same day.

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Stability under test conditions: No analyses were undertaken to assess the stability, homogeneity or achieved concentrations of the test material in the vehicles.

Species:

guinea pig

Strain:

Dunkin-Hartley

Sex:

male/female

Details on test animals and environmental conditions:

TEST ANIMALS
- Females nulliparous and non-pregnant: Not specified
- Age at study initiation: All animals were six to eight weeks of age before treatment on Day 1.
- Weight at study initiation: On Day 1, the bodyweight range of the control animals was 346 - 449 g and of the test animals was 363 - 468 g.
- Housing: Housed in stainless steel cages, with grid floors and tops. The grid floors ensured rapid removal of waste material to undertrays which were cleaned out as necessary. No more than five animals of the same sex were assigned to each cage (animals were allocated to each cage without further selective procedures). The cages were suspended in mobile stainless steel racks.
- Diet: ad libitum access to a commercially available pelleted guinea pig diet.
- Water: ad libitum access to tap water taken from the public supply.
- Acclimation period: An acclimatisation period of at least six days but not more than sixteen days was allowed between arrival at the laboratory and first administration of the test material in the main study. During this period the health of the animals was monitored

ENVIRONMENTAL CONDITIONS
- Temperature: 18 °C (range 15 - 23 °C)
- Humidity: 55 % R.H. (range 40 - 70 %)
- Air changes: 15 air changes per hour without re-circulation.
- Photoperiod: 12 hours of artificial light per day; there was no source of natural light.

Route:

intradermal

Vehicle:

propylene glycol

Concentration / amount:

5 % w/v in propylene glycol and 5 % w/v in propylene glycol in FCA

Day(s)/duration:

Treatment on Day 1

Adequacy of induction:

highest concentration used causing mild-to-moderate skin irritation and well-tolerated systemically

Route:

epicutaneous, occlusive

Vehicle:

propylene glycol

Concentration / amount:

50 % w/v in propylene glycol

Day(s)/duration:

Treatment on Day 8

Adequacy of induction:

highest concentration used causing mild-to-moderate skin irritation and well-tolerated systemically

No.:

#1

Route:

epicutaneous, occlusive

Vehicle:

propylene glycol

Concentration / amount:

0.5 and 3 % w/v in propylene glycol

Day(s)/duration:

Treatment on Day 21

Adequacy of challenge:

highest non-irritant concentration

No. of animals per dose:

10 / sex / group

Details on study design:

RANGE FINDING TESTS:
- The study design included a primary skin irritation screen which imposed limits on the challenge concentration of test material used during the main study. The concentrations used for first and second induction were determined during the previous study.
- Topical Challenge Administration
Three guinea-pigs received a single intradermal injection of 0.1 mL FCA 17 days before topical application to simulate the FCA insult that the main study animals receive before challenge application. The hair was removed from both flanks of the three animals. Topical application of 0.03 mL of four concentrations of test material in the selected challenge vehicle was administered to the four test sites on each guinea-pig.
Each test formulation was applied to a 1 cm diameter absorbent patch (A1-test, Imeco AB, Södertälje, Sweden) which was applied to the skin and covered by an occlusive dressing (Blenderm and Elastoplast) for 24 hours.
Reactions to treatment were assessed approximately 24 and 48 hours after removal of the dressings.

MAIN STUDY
A. INDUCTION EXPOSURE
- Primary induction: The induction procedures were primary induction by intradermal injection on Day 1 and secondary induction by occluded topical application on Day 8. Dermal responses to primary and secondary induction were assessed approximately 24 hours and 48 hours after injection or removal of the occlusive dressings.
Three pairs of injections (0.1 mL) were made deep into the dermis, such that on either side of the dorsal median line there were three injection sites in a row parallel to the spinal column. All injection sites lay near the periphery of a dermal test site 4 x 2 cm long, overlying the scapulae. The anterior and middle sites were positioned close together and distant from the posterior sites.
In the test material treatment group, the anterior sites were injected with FCA, the middle sites with the test material in propylene glycol and the posterior sites with the test material in propylene glycol in FCA. In the control group, the anterior sites were injected with FCA, the middle sites with propylene glycol and the posterior sites with propylene glycol in FCA.
- Secondary induction: On Day 8, the dermal sites overlying the scapulae were treated by topical application of 0.6 mL of 50 % w/v test material in propylene glycol to test animals, while controls received 0.6 mL of the vehicle. Each dose was applied to a 4 x 2.5 cm absorbent patch (Whatman No. 3 filter paper) which was applied to the skin and covered by an occlusive dressing (Blenderm and Elastoplast) for 48 hours. The application site was wiped with a paper tissue moistened with the vehicle immediately after removal of the bandage.
- No. of exposures: 2
- Exposure period: 48 h for secondary induction
- Control group: The control animals (Group 1) were treated identically to the test animals (Group 2) during the induction and challenge procedures, except that during induction the test material was replaced by vehicle.

B. CHALLENGE EXPOSURE
- Site: Both flanks of all animals were clipped on Day 21. On Day 22 these areas were wet shaven to reveal a 5 x 10 cm area on the left flank and a 5 x 10 cm area on the right flank. Approximately three hours later the left site was treated by topical application of 0.03 mL of the vehicle while the right side received 0.03 mL of the maximum non-irritant concentration to one site and a dilution to a second site. The doses were applied to 1 cm diameter absorbent patches (A1-test) and covered by an occlusive dressing for 24 hours. The test site was wiped with a paper tissue moistened with vehicle immediately after removal of the bandage.
- Evaluation: The challenge sites were examined approximately 24 and 48 hours after removal of the occlusive dressings.

OTHER:
- Freunds Complete Adjuvant: Freunds Complete Adjuvant (FCA) was prepared immediately before use by emulsifying equal volumes of purified water and the concentrate of the complete adjuvant. The term Freunds Complete Adjuvant or the abbreviation FCA used throughout this RSS refers to a 1:1 mixture of the adjuvant concentrate with purified water. Fresh doses were prepared on the day of administration and any unused material was disposed of on the same day.
- Preparation of animals: An area of skin 4 cm long and 6 cm wide overlying the scapulae was clipped free of hair with electric clippers on the day before treatment commenced.
- Scoring: The presence or absence of erythema or swelling of the treated skin was assessed without knowledge of the number or group identity of the animal under examination. The degree of reaction was scored on a five point scale:
0 = No response
+ = Barely perceptible erythema
1 = Slight erythema
2 = Moderate erythema
3 = Severe erythema
Other reactions to treatment were recorded on an individual basis (present or absent).
Barely perceptible erythema (Grade +) is often a non-specific response to the dosing procedure and is not considered to be a significant or conclusive indication of delayed contact hypersensitivity.
- Bodyweight: The bodyweight of each animal was recorded at weekly intervals to detect treatment-related depression of growth or individual cases of ill-health.
- Termination: All animals were killed at termination of the study, without necropsy.

Positive control substance(s):

no

Key result

Reading:

1st reading

Hours after challenge:

24

Group:

test chemical

Dose level:

0.5 w/v in propylene glycol

No. with + reactions:

0

Total no. in group:

20

Remarks on result:

no indication of skin sensitisation

Key result

Reading:

1st reading

Hours after challenge:

24

Group:

test chemical

Dose level:

3 % w/v in propylene glycol

No. with + reactions:

0

Total no. in group:

20

Remarks on result:

no indication of skin sensitisation

Reading:

1st reading

Hours after challenge:

24

Group:

test chemical

Dose level:

propylene glycol

No. with + reactions:

0

Total no. in group:

20

Remarks on result:

no indication of skin sensitisation

Reading:

1st reading

Hours after challenge:

24

Group:

negative control

Dose level:

0.5 % w/v in propylene glycol

No. with + reactions:

1

Total no. in group:

20

Remarks on result:

no indication of skin sensitisation

Reading:

1st reading

Hours after challenge:

24

Group:

negative control

Dose level:

3 % w/v in propylene glycol

No. with + reactions:

3

Total no. in group:

20

Remarks on result:

no indication of skin sensitisation

Reading:

1st reading

Hours after challenge:

24

Group:

negative control

Dose level:

propylene glycol

No. with + reactions:

0

Total no. in group:

20

Remarks on result:

no indication of skin sensitisation

Reading:

1st reading

Hours after challenge:

24

Group:

positive control

Remarks on result:

not measured/tested

- Induction: Intradermal administration of 5 % w/v test material in propylene glycol in the adjuvant, caused slight or moderate erythema, pallor and occasional discolouration. Injection of 5 % w/v test material in propylene glycol alone gave rise to a single case of pallor.

Topical application of 50 % w/v test material in propylene glycol caused slight or moderate erythema, oedema, exfoliation and occasional eschar formation, fissuring and loss of elasticity.

 

- Challenge: Challenge application of 3 % w/v test material in propylene glycol caused slight erythema in three control animals and barely perceptible erythema in three test and four control animals. Exfoliation was seen in three test and five control animals. 

Challenge with 0.5 % w/v test material in propylene glycol caused slight erythema in one control animal and barely perceptible erythema in a further control animal; exfoliation was seen in two control animals. No dermal reaction was evident amongst the test animals.

Barely perceptible erythema and exfoliation were seen in two control animals challenged with propylene glycol alone.

 

- General health and bodyweight: The animals remained in overt good health and achieved anticipated overall bodyweight gains. On Day 25, the bodyweight range of the control animals was 503 - 730 g and of the test animals 493 - 695 g.

Interpretation of results:

other: Not classified according to EU criteria.

Conclusions:

Under the conditions of this study, repeated administration of the test material did not cause delayed contact hypersensitivity in the guinea-pig.

Executive summary:

The potential of the test material to cause delayed contact hypersensitivity in guinea-pigs was assessed by the Magnusson-Kligman Maximisation Test in accordance with the standardised guideline OECD 406 and US EPA OPP 81-6 in compliance with GLP.

The closely-clipped dorsa of ten male and ten female Dunkin-Hartley guinea-pigs were subject to intradermal injections of Freunds Complete Adjuvant, 5 % w/v test material in propylene glycol and 5 % w/v test material in propylene glycol in the adjuvant on Day 1. Seven days later the same area of skin was treated by topical application of 50 % w/v test material in propylene glycol and the test site was covered by an occlusive dressing for 48 hours.

The same induction procedures were carried out on a contemporaneous control group of ten male and ten female animals, except that the test material was replaced by vehicle in all doses.

On Day 22, all animals were challenged by occluded application of propylene glycol to the left flank, and 3 % and 0.5 % w/v test material in propylene glycol to two sites on the right flank. The occlusive dressings were removed on the following day and the condition of the test sites was assessed approximately 24 and 48 hours later.

Intradermal administration of 5 % w/v test material in propylene glycol in the adjuvant, caused slight or moderate erythema, pallor and occasional discolouration. Injection of 5 % w/v test material in propylene glycol alone gave rise to a single case of pallor.

Topical application of 50 % w/v test material in propylene glycol caused slight or moderate erythema, oedema, exfoliation and occasional eschar formation, fissuring and loss of elasticity.

Challenge application of 3 % w/v test material in propylene glycol gave rise to a significant response (slight erythema or a more marked reaction) in no test but three control animals. Challenge with 0.5 % w/v test material in propylene glycol gave rise to a significant response in no test but one control animal only. Challenge application of propylene glycol alone caused no significant response in any animal.

Under the conditions of this study, repeated administration of the test material did not cause delayed contact hypersensitivity in the guinea-pig.

Reference 2

Endpoint:

skin sensitisation: in vivo (non-LLNA)

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

01 March 1994 to 25 March 1994

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

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

Qualifier:

according to guideline

Guideline:

EPA OPP 81-6 (Skin Sensitisation)

Deviations:

no

GLP compliance:

yes

Type of study:

guinea pig maximisation test

Justification for non-LLNA method:

Data from existing non-LLNA study available.

Specific details on test material used for the study:

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing: Aqueous preparations correspond to the practical use conditions.
- Preparation of the test material formulations: Immediately before test material application with Ultraturrax or with a magnetic stirrer. Formulations of the test material were prepared gravimetrically; all concentrations were determined in weight/weight.
- Analysis of the test material preparations: The stability of the test material in 0.5 % aqueous CMC preparation over a time period of at least 24 hours was confirmed by analysis. The homogeneity of the test material preparations was provided by stirring.

Species:

guinea pig

Strain:

Pirbright-Hartley

Remarks:

Pirbright White, Dunkin Hartley Crl: (HA)BR (SPF)

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Females nulliparous and non-pregnant: Not specified
- Age at study initiation: Young adult.
- Weight at study initiation: 310 - 360 g
- Housing: Makrolon type IV, with 5 animals per cage.
- Diet: ad libitum
- Water: Tap water ad libitum; about 2 g of ascorbic acid per 10 L water was added to the drinking water twice a week.
- Acclimation period: 7 days before the beginning of the study.

ENVIRONMENTAL CONDITIONS
- Temperature: 20 - 24 °C
- Humidity: 30 - 70 %
- Photoperiod: 12 h light (06.00 to 18.00) : 12 h darkness (18.00 to 06.00)

Route:

intradermal

Vehicle:

other:

Remarks:

0.5 % solution of Tylose® CB 30 000 in aqua bidest., or in Freund's adjuvant / 0.9 % aqueous NaCl solution (1:1), or 0.5 % solution of Tylose® CB 30 000 in aqua bidest.

Concentration / amount:

5 %

Day(s)/duration:

24 h

Adequacy of induction:

highest concentration used causing mild-to-moderate skin irritation and well-tolerated systemically

Route:

epicutaneous, occlusive

Vehicle:

other:

Remarks:

0.5 % solution of Tylose® CB 30 000 in aqua bidest., or 0.5 % solution of Tylose® CB 30 000 in aqua bidest.

Concentration / amount:

5 %

Day(s)/duration:

48 h

Adequacy of induction:

highest concentration used causing mild-to-moderate skin irritation and well-tolerated systemically

No.:

#1

Route:

epicutaneous, occlusive

Vehicle:

other:

Remarks:

0.5 % solution of Tylose® CB 30 000 in aqua bidest., or 0.5 % solution of Tylose® CB 30 000 in aqua bidest.

Concentration / amount:

1 %

Day(s)/duration:

24 hours

Adequacy of challenge:

highest non-irritant concentration

No. of animals per dose:

20

Details on study design:

RANGE FINDING TESTS:
- Pre-test: For detecting a possible influence on irritating effects of previous intradermal treatment with Freund's adjuvant, animals pre-treated with Freund's adjuvant / 0.9 % aqueous NaCl-solution (1:1) each, in the same manner as intradermal induction about 3 or 4 weeks prior to the application of the test material were used.
In the preliminary test after two 24-hour percutaneous occlusive applications within 96 hours, the minimum irritant concentration was found to be a 2 % test material preparation in 0.5 % solution of Tylose® CB 30 000 in aqua bidest. The maximum non-irritant concentration was found to be a 1 % test material preparation in 0.5 % solution of Tylose® CB 30 000 in aqua bidest. (48 hours after the beginning of application).
- Applicability: It was possible to inject a 5 % test material preparation in 0.5 % solution of Tylose® CB 30 000 in aqua bidest. resp. in Freund's adjuvant / 0.9 % aqueous NaCl-solution (1:1) with a syringe.
- Amount applied: 2 x 2 cm filter paper strips were applied to the skin of the flanks under an occlusive dressing (the bandage consists of rubberised linen patches 4 x 4 cm from Russka, test patch 5 x 5 cm of Idealbinde from Pfälzische Verbandstoff-Fabrik, and Fixomull® Stretch (adhesive fleece) from Beiersdorf AG). In the case of liquid test material formulations the test filter paper strip was soaked in the test material formulation; in the case of solid test material formulations the paper strip was coated with an approx. 0.5 mm thick layer of the test material formulation; thus, the animals were exposed to about 0.15 g of the test material formulation.
- Exposure period: The test material was applied 2 times for 24 hours within a period of 96 hours in order to detect non-specific phenomena that are not caused by a sensitisation reaction but could possibly be attributed to a shift in the irritation threshold.
- Site of application: Flank
- Number of test animals: 4 or 8 per test concentration.
- Readings: About 24 and 48 h after the beginning of application.

MAIN STUDY
A. INDUCTION EXPOSURE: Intradermal Induction
- No. of exposures: One; 6 intradermal injections in groups of 2 per animal
- Exposure period: 24 h
Front row: 2 injections each of 0.1 mL Freund's adjuvant without test material emulsified with 0.9 % aqueous NaCl solution in a ratio of 1:1.
Middle row: 2 injections each of 0.1 mL of the test material formulation.
Back row: 2 injections each of 0.1 mL Freund's adjuvant / 0.9 % aqueous NaCl solution (1:1) with test material.
- Control group: 20 animals (two groups of 10)
The animals were given the same injections as the test material group but without the test material, only with the formulating agent.
- Site: Shoulder
- Readings: 24 h after the beginning of application.
- Concentrations: 5 % solution

INDUCTION EXPOSURE: Percutaneous induction was carried out one week after intradermal induction.
- No. of exposures: One
- Exposure period: 48 h
- Control group: The control groups were treated analogously to the test group but only with the vehicle without the test material.
- Site: Shoulder, same area as in the case of the previous intradermal application.
- Readings: 48 h after the beginning of application.
- Concentrations: 5 % solution
2 x 4 cm filter paper strips were applied to the skin of the shoulder under an occlusive dressing (the bandage consists of rubberised linen patches 4 x 6 cm from Russka and Fixomull® Stretch (adhesive fleece) from Beiersdorf AG). The filter paper strip was soaked in the test material formulation. Thus, the animals were exposed to about 0.3 g of the test material formulation.

B. CHALLENGE EXPOSURE
- No. of exposures: One
- Day(s) of challenge: 21 days after intradermal induction.
- Exposure period: 24 h
- Test groups: Treatment of the test group with the test material formulation. Additionally 0.5 % solution of Tylose® CB 30 000 in aqua bidest. was applied as vehicle.
- Control group: Treatment of control group 1 with the test material formulation. Additionally 0.5 % solution of Tylose® CB 30 000 in aqua bidest. was applied as vehicle. Control group 2 only received 0.5 % solution of Tylose® CB 30 000 in aqua bidest.
- Site: Intact flank.
- Concentrations: 2 x 2 cm filter paper strips were applied to the skin of the flank under an occlusive dressing (the bandage consists of rubberised linen patches 4 x 4 cm from Russka, test patch 5 x 5 cm of Idealbinde from Pfälzische Verbandstoff-Fabrik, and Fixomull® Stretch (adhesive fleece) from Beiersdorf AG). The test filter paper strip was soaked in the test material formulation. Thus the animals were exposed to about 0.15 g of the test material formulation.
- Readings: 24 and 48 h after the removal of the patch.

OTHER:
- Randomisation: According to Salfi, R.: A Long Period Random Number Generator with Application to Permutation. Compstat 1974, pp. 28 - 35.
- Bodyweight: Weight check of the individual animals: At the beginning of the study (day 0) and at the end of the study (last day of observation).
- Clipping: Clipping of the test animals about 3 hours before each test material application at the appropriate application sites.
- General observations: A check was made twice each workday and once on Saturdays, Sundays and on public holidays for general observations and for any dead or moribund animal.
- Assessment of skin findings: According to Draize, J.H. (1959): Appraisal of the Safety of Chemicals in Foods, Drugs and Cosmetics. The Association of Food and Drug Officials of the United States Austin, Texas):
Erythema and eschar formation
0 = No erythema
1 = Very slight erythema (barely perceptible)
2 = Well-defined erythema
3 = Moderate to severe erythema
4 = Severe erythema (beet redness) to slight eschar formation (injuries in depth)

Oedema formation
0 = No oedema
1 = Very slight oedema (barely perceptible)
2 = Slight oedema (edges of area well defined by definite raising)
3 = Moderate oedema (raised approximately 1 mm)
4 = Severe oedema (raised more than 1 mm and extending beyond the area of exposure)

Positive control substance(s):

yes

Remarks:

A positive control (reliability check) with a known sensitiser (1-chloro-2,4-dinitrobenzene) is not included in this study however a separate study is performed twice a year in the laboratory and the most recent results included.

Positive control results:

The positive control with 1-chloro-2,4-dinitrobenzene showed that the test system was able to detect sensitising compounds under the laboratory conditions chosen.

Reading:

1st reading

Hours after challenge:

24

Group:

test chemical

Dose level:

0.5 % solution of Tylose® CB 30 000 in aqua bidest

No. with + reactions:

0

Total no. in group:

20

Remarks on result:

no indication of skin sensitisation

Reading:

1st reading

Hours after challenge:

24

Group:

test chemical

Dose level:

1 % in 0.5 % solution of Tylose® CB 30 000 in aqua bidest

No. with + reactions:

0

Total no. in group:

20

Remarks on result:

no indication of skin sensitisation

Reading:

1st reading

Hours after challenge:

24

Group:

other: control group 1

Dose level:

1 % in 0.5 % solution of Tylose® CB 30 000 in aqua bidest

No. with + reactions:

0

Total no. in group:

10

Remarks on result:

no indication of skin sensitisation

Reading:

1st reading

Hours after challenge:

24

Group:

other: control group 1

Dose level:

0.5 % solution of Tylose® CB 30 000 in aqua bidest

No. with + reactions:

0

Total no. in group:

10

Remarks on result:

no indication of skin sensitisation

Reading:

1st reading

Hours after challenge:

24

Group:

other: control group 2

Dose level:

0.5 % solution of Tylose® CB 30 000 in aqua bidest

No. with + reactions:

0

Total no. in group:

10

Remarks on result:

no indication of skin sensitisation

After intradermal induction well-defined erythema and slight oedema could be observed at the injection sites of the control and test group animals at which only Freund's adjuvant / 0.9 % aqueous NaCl-solution (1:1) was applied. Injection of a 5 % test material preparation in 0.5 % solution of Tylose® CB 30 000 in aqua bidest. caused well-defined erythema and slight oedema in the test group animals. The 5 % test material preparation in Freund's adjuvant / 0.9 % aqueous NaCl solution (1:1) caused well-defined erythema and slight oedema in about half of the injection sites of the test group animals. The other injection sites exhibited necrotic skin changes and slight oedema. The control group animals injected with the vehicle did not show any skin reactions.

After the percutaneous induction with a 5 % test material preparation in 0.5 % solution of Tylose® CB 30 000 in aqua bidest. well-defined erythema and slight oedema in addition to incrustation, partially open (caused by the intradermal induction) could be observed in the test group animals. The control group animals treated with the vehicle alone showed the same skin reactions.

The challenge with the 1 % test material preparation in 0.5 % solution of Tylose® CB 30 000 in aqua bidest. did not cause any skin reactions neither in the test group animals nor in the animals of control group 1.

0.5 % solution of Tylose® CB 30 000 in aqua bidest. applied as vehicle did not cause any skin reactions in the animals of the control groups and the test group.

The number of animals sensitised is primarily taken into account in the evaluation. The control animals are used to rule out a substance-induced primary skin irritation.

The findings obtained 24 hours after the removal of the patch are taken into account for the determination of the sensitisation rate.

Interpretation of results:

other: Not classified according to EU criteria.

Conclusions:

Under the conditions of the study the test material does not have a sensitising effect on the skin of the guinea pig in the Maximisation Test.

Executive summary:

The skin sensitisation potential of the test material was assessed in accordance with the standardised guidelines OECD 406, EU Method B.6. and US EPA OPP 81-6 in compliance with GLP.

The test material was tested for its sensitising effect on the skin of the guinea pig in the Maximisation Test based on the method of Magnusson and Kligman. After intradermal induction with 5 % test material preparations, slight to well-defined signs of irritation and in some cases necrotic skin changes could be observed in the test group animals. Percutaneous induction with a 5 % test material preparation led to incrustation, partially open (caused by the intradermal induction) in addition to well-defined erythema and slight oedema in the test group animals. 

The number of animals with skin findings after the challenge (21 days after intradermal induction) is summarised below. Results are shown as the number of positive reactions/number of animals tested; readings were carried out 24 h after the removal of the patch.

- Test group

1 % test material in 0.5 % solution of Tylose® CB 30 000 in aqua bidest: 0 / 10

0.5 % solution of Tylose® CB 30 000 in aqua bidest: 0 / 10

- Control group 1

1 % test material in 0.5 % solution of Tylose® CB 30 000 in aqua bidest: 0 / 10

0.5 % solution of Tylose® CB 30 000 in aqua bidest: 0 / 10

- Control group 2

This group, that had been intended for a potential 2nd challenge, was not treated with the test material, since a 2nd challenge was not necessary on the basis of the unambiguous results of the 1st challenge.

0.5 % solution of Tylose® CB 30 000 in aqua bidest: 0 / 10

Under the conditions of the study it was concluded that the test material does not have a sensitising effect on the skin of the guinea pig in the Maximisation Test.

Reference 3

Endpoint:

skin sensitisation: in vitro

Data waiving:

study scientifically not necessary / other information available

Justification for data waiving:

an in vitro skin sensitisation study does not need to be conducted because adequate data from an in vivo skin sensitisation study are available

Justification for type of information:

An in vitro skin sensitisation study does not need to be conducted because adequate data from in vivo skin sensitisation studies are available.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not sensitising)

Additional information:

Skin Sensitisation In Vivo: Rees (1995)

The potential of the test material to cause delayed contact hypersensitivity in guinea-pigs was assessed by the Magnusson-Kligman Maximisation Test in accordance with the standardised guideline OECD 406 and US EPA OPP 81-6 in compliance with GLP. The study was awarded a reliability score of 1 in accordance with the criteria set forth by Klimisch et al. (1997).

The closely-clipped dorsa of ten male and ten female Dunkin-Hartley guinea-pigs were subject to intradermal injections of Freunds Complete Adjuvant, 5 % w/v test material in propylene glycol and 5 % w/v test material in propylene glycol in the adjuvant on Day 1. Seven days later the same area of skin was treated by topical application of 50 % w/v test material in propylene glycol and the test site was covered by an occlusive dressing for 48 hours.

The same induction procedures were carried out on a contemporaneous control group of ten male and ten female animals, except that the test material was replaced by vehicle in all doses.

On Day 22, all animals were challenged by occluded application of propylene glycol to the left flank, and 3 % and 0.5 % w/v test material in propylene glycol to two sites on the right flank. The occlusive dressings were removed on the following day and the condition of the test sites was assessed approximately 24 and 48 hours later.

Intradermal administration of 5 % w/v test material in propylene glycol in the adjuvant, caused slight or moderate erythema, pallor and occasional discolouration. Injection of 5 % w/v test material in propylene glycol alone gave rise to a single case of pallor.

Topical application of 50 % w/v test material in propylene glycol caused slight or moderate erythema, oedema, exfoliation and occasional eschar formation, fissuring and loss of elasticity.

Challenge application of 3 % w/v test material in propylene glycol gave rise to a significant response (slight erythema or a more marked reaction) in no test but three control animals. Challenge with 0.5 % w/v test material in propylene glycol gave rise to a significant response in no test but one control animal only. Challenge application of propylene glycol alone caused no significant response in any animal.

Under the conditions of this study, repeated administration of the test material did not cause delayed contact hypersensitivity in the guinea-pig.

Supporting Study: Rossbacher & Hellwig (1995)

The skin sensitisation potential of the test material was assessed in accordance with the standardised guidelines OECD 406, EU Method B.6. and US EPA OPP 81-6 in compliance with GLP. The study was awarded a reliability score of1in accordance with the criteria set forth by Klimisch et al. (1997).

The test material was tested for its sensitising effect on the skin of the guinea pig in the Maximisation Test based on the method of Magnusson and Kligman. After intradermal induction with 5 % test material preparations, slight to well-defined signs of irritation and in some cases necrotic skin changes could be observed in the test group animals. Percutaneous induction with a 5 % test material preparation led to incrustation, partially open (caused by the intradermal induction) in addition to well-defined erythema and slight oedema in the test group animals. 

The number of animals with skin findings after the challenge (21 days after intradermal induction) is summarised below. Results are shown as the number of positive reactions/number of animals tested; readings were carried out 24 h after the removal of the patch.

- Test group

1 % test material in 0.5 % solution of Tylose® CB 30 000 in aqua bidest: 0 / 10

0.5 % solution of Tylose® CB 30 000 in aqua bidest: 0 / 10

- Control group 1

1 % test material in 0.5 % solution of Tylose® CB 30 000 in aqua bidest: 0 / 10

0.5 % solution of Tylose® CB 30 000 in aqua bidest: 0 / 10

- Control group 2

This group, that had been intended for a potential 2nd challenge, was not treated with the test material, since a 2nd challenge was not necessary on the basis of the unambiguous results of the 1st challenge.

0.5 % solution of Tylose® CB 30 000 in aqua bidest: 0 / 10

Under the conditions of the study it was concluded that the test material does not have a sensitising effect on the skin of the guinea pig in the Maximisation Test.

Justification for classification or non-classification

In accordance with the criteria for classification as defined in Annex I, Regulation (EC) No 1272/2008, the substance does not require classification with respect to skin sensitisation.