Registration Dossier

Administrative data

Description of key information

An Open Epicutaneous Test (OET) was performed on guinea pigs to assess the skin sensitization potential of test chemical. The Pretest was performed to establish the primary irritating threshold concentration of test substance. This test was performed by applying with a pipette 0.025 ml of each concentration to skin areas measuring 2 cm2. The reactions were read after 24, 48 and/or 72h. It was observed that none of the guinea pigs induced contact sensitization at challenge concentration of 8%.Thus, the test chemical was considered to be not sensitizing on skin of guinea pigs at concentration of 8% in an Open Epicutaneous Test (OET).

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
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
data from handbook or collection of data
Justification for type of information:
data is from peer reviewed journals
Qualifier:
according to guideline
Guideline:
other: Open Epicutaneous test
Principles of method if other than guideline:
An Open Epicutaneous Test (OET) was performed on guinea pigs to assess the skin sensitization potential of test chemical.
GLP compliance:
not specified
Type of study:
other: Open Epicutaneous Test (OET)
Justification for non-LLNA method:
not specified
Species:
guinea pig
Strain:
not specified
Sex:
male/female
Details on test animals and environmental conditions:
- Weight at study initiation: 300-450g
Route:
epicutaneous, open
Vehicle:
other: ethanol, acetone, H20, petroleum, PEG and/or other suitable vehicles
Concentration / amount:
Concentration:8%
Amount: 0.1ml
Day(s)/duration:
3 weeks (0-21 days)
Adequacy of induction:
not specified
No.:
#1
Route:
epicutaneous, open
Vehicle:
other: ethanol, acetone, H20, petroleum, PEG and/or other suitable vehicles
Concentration / amount:
8% - 0.025 ml
Day(s)/duration:
72 hours
Adequacy of challenge:
not specified
No. of animals per dose:
6–8 male and female guinea pigs
Details on study design:
RANGE FINDING TESTS: The Pretest was performed to establish the primary irritating threshold concentration of test substance. For this, a single application of 0.025 ml of each test concentration (e.g, 100, 30, 10 and 3%) is simultaneously performed on one of the areas measuring 2 cm2 of the flank skin previously clipped and marked with a circular stamp. Reactions are read 24 h after the application of the test material. On the basis of preliminary study, the maximal non-irritating concentration selected was 8%. Thus the sensitization test was conducted at a dose of 8% of test chemical.

MAIN STUDY
A. INDUCTION EXPOSURE
- No. of exposures: 21
- Exposure period:24 hours
- Test groups:6-8 guinea pigs
- Control group:3 guinea pigs
- Site: an area measuring 8 cm2 on the clipped flank skin of the guinea pigs
- Frequency of applications: The applications are repeated daily for 3 weeks or done 5 times weekly during 4 weeks
- Duration: 21 days (3 weeks)
- Concentrations: 8% in vehicle.
Amount: 0.1ml

B. CHALLENGE EXPOSURE
- No. of exposures: 2
- Day(s) of challenge: on days 21 and 35
- Exposure period:24 hours
- Test groups: 6-8 guinea pigs
- Control group: 3 guinea pigs
- Site: contralateral flank measuring 2 cm2
- Concentrations: Concentration:8%
Amount: 0.025ml
- Evaluation (hr after challenge): 24,48 and/or 72h.
Challenge controls:
The 10 controls were either left untreated or treated with 0.1 ml aliquot of the vehicle for 21 days
Positive control substance(s):
not specified
Positive control results:
no data available
Key result
Reading:
1st reading
Hours after challenge:
72
Group:
test chemical
Dose level:
8%
No. with + reactions:
0
Total no. in group:
6
Clinical observations:
no signs of dermal sensitization pbserved
Remarks on result:
no indication of skin sensitisation

Table: Results of the sensitization study

Preparation

Maximum user concentration

Sensitization procedure

Human

Guinea pigs

Concentration%

HMT or RPT result

OET result

test chemical

1.5

8

-

-

 

Interpretation of results:
other: not sensitizing
Conclusions:
The test chemical was considered to be not sensitizing on skin of guinea pigs at concentration of 8% in an Open Epicutaneous Test (OET).
Executive summary:

An Open Epicutaneous Test (OET) was performed on guinea pigs to assess the skin sensitization potential of test chemical.

 

The Pretest was performed to establish the primary irritating threshold concentration of test substance. For this, a single application of 0.025 ml of each test concentration (e.g, 100, 30, 10 and 3%) is simultaneously performed on one of the areas measuring 2 cm2 of the flank skin previously clipped and marked with a circular stamp. Reactions are read 24 h after the application of the test material. On the basis of preliminary study, the maximal non-irritating concentration selected was 8%. Thus the sensitization test was conducted at a dose of 8% of test chemical.

 

On day 1 during induction, 0.1 ml of test chemical was applied at concentrations of 8% in vehicle to an area measuring 8 cm2 on the clipped flank skin of the guinea pigs. The applications are repeated daily for 3 weeks or done 5 times weekly during 4 weeks, usually on the same skin sites. The application sites were left uncovered and the reactions, if continuous daily applications were performed, can be read 24 h after each application, or at the end of each week. 

To determine whether or not contact sensitization was induced, all groups of guinea pigs previously treated for 21 days, as well as 10 untreated, or only pretreated with the vehicle, controls are tested on days 21 and 35 on the contralateral flank with the test material. This test was performed by applying with a pipette 0.025 ml of each concentration to skin areas measuring 2 cm2. The reactions were read after 24, 48 and/or 72h.

 

It was observed that none of the guinea pigs induced contact sensitization at challenge concentration of 8%.Thus, the test chemical was considered to be not sensitizing on skin of guinea pigs at concentration of 8% in an Open Epicutaneous Test (OET).

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (not sensitising)
Additional information:

Various studies were performed on humans, guinea pigs to assess the dermal sensitization potential of Test chemical. The studies are summarized as follows:

 

An Open Epicutaneous Test (OET) was performed on guinea pigs to assess the skin sensitization potential of test chemical. The Pretest was performed to establish the primary irritating threshold concentration of test substance. For this, a single application of 0.025 ml of each test concentration (e.g, 100, 30, 10 and 3%) is simultaneously performed on one of the areas measuring 2 cm2 of the flank skin previously clipped and marked with a circular stamp. Reactions are read 24 h after the application of the test material. On the basis of preliminary study, the maximal non-irritating concentration selected was 8%. Thus the sensitization test was conducted at a dose of 8% of test chemical. On day 1 during induction, 0.1 ml of test chemical was applied at concentrations of 8% in vehicle to an area measuring 8 cm2 on the clipped flank skin of the guinea pigs. The applications are repeated daily for 3 weeks or done 5 times weekly during 4 weeks, usually on the same skin sites. The application sites were left uncovered and the reactions, if continuous daily applications were performed, can be read 24 h after each application, or at the end of each week.  To determine whether or not contact sensitization was induced, all groups of guinea pigs previously treated for 21 days, as well as 10 untreated, or only pretreated with the vehicle, controls are tested on days 21 and 35 on the contralateral flank with the test material. This test was performed by applying with a pipette 0.025 ml of each concentration to skin areas measuring 2 cm2. The reactions were read after 24, 48 and/or 72h. It was observed that none of the guinea pigs induced contact sensitization at challenge concentration of 8%.Thus, the test chemical was considered to be not sensitizing on skin of guinea pigs at concentration of 8% in an Open Epicutaneous Test (OET).

 

This result is supported by the study conducted on humans for test chemical. The test chemical was evaluated in Kligman maximization test conducted on 25 healthy adult volunteers. The test substance was applied under occlusion to the same site on the forearms of all subjects for 5 alternate day 48 h periods. Patch sites were pre-treated for 24 h with 1 ml of 5% aqueous sodium lauryl sulfate (SLS) under occlusion. Following a 10 day rest period, a challenge patch of 8% test material was applied to a fresh site on the scapular back for 48 h under occlusion. Prior to challenge, 10% SLS was applied to the test site for 1 h before application of ionone. The challenge site was read at patch removal and again on each of two successive days. No sensitization reactions were reported after challenge exposure. It was considered that test chemical does not cause skin sensitization in humans.

 

These studies were further supported by the results obtained for test chemical by using Guinea pig maximization assay. Induction and challenge were conducted with 10% of test chemical. No sensitization reactions were reported after challenge exposure. It was concluded that the test substance does not cause skin sensitization in guinea pigs.

 

 

 FREUND'S COMPLETE ADJUVANT (FCAT) test was conducted in guinea pigs (male and female) to determine the skin sensitization potential of test chemical. Male and female guinea pigs, weighing 300-450g, are used. Two groups of 10-20 guinea pigs each are involved: (a) the experimental group, and (b) the control group. Additionally, 4 guinea pigs are used for determination of the primary irritating concentration of the test substance so that nonirritating concentrations may be chosen for challenge. Shortly prior to starting the treatment, the scapula region of test and control animals was shaved. During days 1-9 the animals of the experimental group are treated with 0.1 ml of the test material in FCA intradermally into the scapula region three times (days I, 5 and 9) on an area of 6 x 2 cm. The animals of the control group are treated with 0.1 ml of FCA only. 1 day before the challenge, a group of 4 guinea pigs is used for the estimation of the threshold irritating concentration after a single topical application by simultaneous application of four different concentrations, e.g. 100, 30, 10 and 3%, to the left flank skin. 0,025 ml of each test concentration is homogeneously spread on 1 of 4 areas measuring 2 cm2 on the flank skin previously clipped and marked with a circular stamp. The minimal irritating concentration was defined as the lowest one causing visible erythema. The maximal nonirritating concentration was defined as the highest one not causing macroscopic reactions in any animal. To determine whether or not contact allergy was induced, both animal groups, the experimental and the control one, are challenged on days 21 and 35 on the contralateral flanks with the same compound at the minimal irritating and some lower nonirritating concentrations. The tests are performed by applying with a pipette 0.025 ml of each concentration to skin areas measuring 2 cm2. The dermal reactions were read 24, 48 and 72 h after application. No signs of dermal irritation or sensitization were observed 72 hours after challenge exposure. The test chemical was considered to be not sensitizing on guinea pigs in FREUND'S COMPLETE ADJUVANT (FCAT) test.

 

Draize test was also performed to evaluate the dermal sensitization potential of Test chemical in guinea pigs. Groups of 6–8 male and female outbred Himalayan white-spotted guinea pigs were used for the study. A dose of 0.05 ml of 0.1% solution of Test chemical in isotonic saline was injected intradermally on day 0 and further doses of 0.1 ml each were injected on 9 alternate days. The total dose injected was 0.95 mg. The treated animals and untreated controls were challenged intradermally with 0.05 ml of a 0.1 per cent solution on days 35 and 49.The evaluation criterion was the mean diameter of the papular reactions at the test sites. No sensitization reactions were observed on the skin of guinea pigs. Hence, the Test chemical was considered to be not sensitizing to skin.

 

 

In an another Guinea Pig Maximization test, groups of 6–8 male and female outbred Himalayan white-spotted guinea pigs were used for the study. On day 0, the animals were injected intradermally with 0.1 ml of a 5% solution of Test chemical, 0.1 ml of a 5% emulsion of ionone in Freund’s complete adjuvant (FCA) and 0.1 ml of FCA alone. Each injection was given twice. In addition, 250 mg of Test chemical dissolved in petrolatum at a concentration of 25% was applied on day 8 to a clipped area of the neck and was kept under occlusive bandage for 48 h. On day 21, Test chemical at a sub-irritant concentration in petrolatum was applied to the flank for 24 h under occlusion. Reactions were read at 24 and 48 h after removal of the patch. No sensitization reactions were observed on the skin of guinea pigs. Hence, the Test chemical was considered to be not sensitizing to skin.

 

A patch test was performed on 179 patients (144 females, 34 male, no sex recorded for 1 patient) suspected of cosmetic allergy. The patients were patch tested with a series of 16 fragrance materials and 9 preservatives. 56 patients suffered, or had suffered, based on history of atopic diseases. For patch testing Silver Patch testers were used. The dermal reactions were evaluated after 48 and 72 hours and scored according to internationally accepted criteria. No dermal reactions were observed on the skin of human volunteers when tested with 10% Test chemical in petrolatum. Hence, Test chemical was considered to be not sensitizing to skin.

 

All these studies lead to a conclusion that Test chemical is indeed not sensitizing to skin. Hence, comparing the above annotations with the criteria of CLP regulation, Test chemical can be classified under the category “Not Classified”.

Respiratory sensitisation

Endpoint conclusion
Endpoint conclusion:
no study available

Justification for classification or non-classification

The skin sensitization potential of test substance were observed in various studies. From the results obtained from these studies it is concluded that the chemical is not likely to cause skin sensitization and hence can be classified as non-skin sensitizer. Hence, comparing the above annotations with the criteria of CLP regulation, Test chemical can be classified under the category “Not Classified”.