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Diss Factsheets

Administrative data

Description of key information

The substance is considered to be not sensitizing to the skin of guinea pigs and humans.

Key value for chemical safety assessment

Skin sensitisation

Link to relevant study records

Referenceopen allclose all

Endpoint:
skin sensitisation: in vitro
Data waiving:
study scientifically not necessary / other information available
Justification for data waiving:
other:
Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
experimental study
Adequacy of study:
weight of evidence
Study period:
1979
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
comparable to guideline study with acceptable restrictions
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 406 (Skin Sensitisation)
Version / remarks:
Open Epicutaneous Test
Deviations:
not applicable
Principles of method if other than guideline:
Modified non-occlusive Draize skin sensitsation test with one ocassion for induction (4 intradermal injections of 0.1 mL aliquots at 2.5-fold concentration of ICC)
GLP compliance:
no
Type of study:
Draize test
Justification for non-LLNA method:
Test performed 1979.
Species:
guinea pig
Strain:
not specified
Sex:
not specified
Details on test animals and environmental conditions:
Not specified.
Route:
epicutaneous, open
Vehicle:
not specified
Concentration / amount:
100, 30, 10, 3 and 1% or lower
Day(s)/duration:
Not specified
Adequacy of induction:
not specified
No.:
#1
Route:
epicutaneous, open
Vehicle:
not specified
Concentration / amount:
Not specified
Day(s)/duration:
Not specified
Adequacy of challenge:
not specified
No. of animals per dose:
Not specified.
Details on study design:
PRELIMINARY STUDY
- Type of study: Irritation test
- Obejctive: To determine the animal irritating concentration and to derive the maximal non-irritant concentration (the highest concentration not inducing macroscopic signs of irritation in any of the animals in the specific test group) as baseline data for the main test
- Concentrations: 4 different concentrations
- Vehicle: Not specified
- Test groups: 1 group of test animals per concentration
- Application site: Left flank skin, skin clipped
- Occlusion: None (non-occlusive)
- Exposure: 24 hours
- Reading: After 24 hours

MAIN TEST
- Phases of the test procedure: Induction (21 daily open applications), challenge (on days 21 and 35)
- Concentrations: 100, 30, 10, 3 and 1% or lower
- Vehicle: Ethanol, acetone, water, vaseline, PEG and/or other suitable vehicles; not further specified
- Application site: Left flank skin, skin clipped
- Occlusion: None (non-occlusive)
- Exposure: Not specified
- Reading: Not specified
Challenge controls:
Yes, one control group with 8 test animals.
Positive control substance(s):
no
Positive control results:
Not applicable.
Group:
negative control
Remarks on result:
no indication of skin sensitisation
Group:
positive control
Remarks on result:
not measured/tested
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
test chemical
Dose level:
6%
No. with + reactions:
0
Total no. in group:
6
Clinical observations:
Not specified
Remarks on result:
no indication of skin sensitisation
Key result
Reading:
1st reading
Hours after challenge:
48
Group:
test chemical
Dose level:
6%
No. with + reactions:
0
Total no. in group:
6
Clinical observations:
Not specified
Remarks on result:
no indication of skin sensitisation
Key result
Reading:
1st reading
Hours after challenge:
72
Group:
test chemical
Dose level:
6%
No. with + reactions:
0
Total no. in group:
6
Clinical observations:
Not specified
Remarks on result:
no indication of skin sensitisation

The results of the Open Epicutaneous Test (OET) were compared with those of the Kligman Human Maximization Test (HMT) by RIFM. The results of the OET coincided with those of the HMT: At a test concentration of 6.0% (purity of the test material >= 97%) no signs skin sensitisation was oberved.

Interpretation of results:
GHS criteria not met
Conclusions:
The test item has no skin sensitising potential in an OET with guinea pigs.
Executive summary:

An open epicutaneous test was conducted in guinea pigs. Induction consisted of 21 daily open applications to the shaved flank of 6-8 guinea pigs per group. One to six experimental and one control group was used. Open challenge applications were made on days 21 and 35. Reactions were read at 24, 48 and 72 hours. No further details were provided.

Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
experimental study
Adequacy of study:
weight of evidence
Study period:
1977-09-14
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
comparable to guideline study with acceptable restrictions
Remarks:
Method applied similar to OECD 406 (modified Draize Test)
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 406 (Skin Sensitisation)
Version / remarks:
Modified Draize Test
Deviations:
not applicable
Principles of method if other than guideline:
- Principle of test: Draize skin sensitsation test with one ocassion for induction (4 intradermal injections of 0.1 mL aliquots at 2.5-fold concentration of ICC)
GLP compliance:
no
Type of study:
Draize test
Justification for non-LLNA method:
Study performed 1978.
Species:
guinea pig
Strain:
Hartley
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Bred in colony of the test facility
- Sex: 4 males and 6 females or vis versa
- Females nulliparous and non-pregnant: not specified
- Weight at study initiation: ca. 350 g
- Housing: Wire mesh cages
- Diet: pelleted guinea pig diet, cabbage and hay ad libitium
- Water: Ad libitium
- Acclimation period: Not specified
Route:
intradermal
Vehicle:
not specified
Concentration / amount:
0.625%
Day(s)/duration:
14 days
Adequacy of induction:
highest concentration used causing mild-to-moderate skin irritation and well-tolerated systemically
No.:
#1
Route:
intradermal and epicutaneous
Vehicle:
not specified
Concentration / amount:
0.25% / 0.1 mL (ICC)
10% / 0.1 mL (ACC)
Day(s)/duration:
24 hours
Adequacy of challenge:
highest non-irritant concentration
No.:
#2
Route:
intradermal and epicutaneous
Vehicle:
not specified
Concentration / amount:
0.25% /0.1 mL (ICC)
10% / 0.1 mL (ACC)
Day(s)/duration:
24 hours
Adequacy of challenge:
highest non-irritant concentration
No. of animals per dose:
10 animals for one single dose-level
Details on study design:
RANGE FINDING TESTS:
Albino guinea pigs of the Hartley strain were treated by intradermal injection to induce sensitization. Two weeks later they were challenged both by intradermal injection and topical application. The procedures were repeated when there were no indications of sensitization.

MAIN STUDY
A. INDUCTION EXPOSURE
- No. of exposures: 1 (equivalent total dose was administered on one occasion as 4 intradermal injections, each 2.5 times the ICC) per Induction (2 in test)
- Exposure period: First and second Induction, 14 days
- Test groups: One test group of ten animals treated with the 2.5-fold of the ICC
- Site: 4 sites overlaying auxillary (2 sites) and inguinal (2 sites) lymph nodes
- Frequency of applications: One occasion per induction
- Concentrations: ICC based on preliminary test, 2.5-fold of ICC
B. CHALLENGE EXPOSURE
- No. of exposures: 1 per challenge (2 challenges in test); 1 x intradermal and 1 x topical application
- Day of challenge: 14 days after induction (day 14)
- Exposure period: 24 hours
- Site: Flanks (shaved)
- Concentrations: ICC and ACC based on preliminary test
- Evaluation: 24 hours after start of challenge period
C. RECHALLENGE EXPOSURE
- Day of rechallenge: 7 days after second challenge (day 42)
- Exposure period: 24 hours
- Test groups: One group of 10 test animals for ICC and ACC
- Site: Flanks (shaved)
- Concentrations: ICC and ACC based on preliminary test
- Evaluation: 24 hours after challenge
- Control group: 4 previously untreated test animals, treated with test substance at the ICC and ACC (0.1 mL aliquots)

SCORING
- Visualisation of positive reactions: with a Philips colour-matching unit with 3 Philips 40 W Actinic Blue 05 fluorescent tubes and 3 Philips 40 W White 35 fluorescent tubes
- Scoring System: Injection reaction, erythema and oedema (2 largest diameters); application reactions scored on a 0 to +++ scale
- Criteria for positive reaction: If the total score of the test groups were significantly greater than the average total score of the control; individual reactions considered positive if score + or greater and there were no erythema reactions in controls
Challenge controls:
Negative controls were introduced in confirming rechallenge. Four previously untreated test animals of the same sex and of similar weight to the test animals treated with the test item.
Positive control substance(s):
no
Positive control results:
Not applicable
Group:
negative control
Remarks on result:
no indication of skin sensitisation
Group:
positive control
Remarks on result:
not measured/tested
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
test chemical
Dose level:
0.25% (ICC) and 10% (ACC)
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
Not specified
Remarks on result:
no indication of skin sensitisation
Key result
Reading:
2nd reading
Hours after challenge:
24
Group:
test chemical
Dose level:
ICC and ACC
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
Not specified
Remarks on result:
no indication of skin sensitisation
Interpretation of results:
GHS criteria not met
Conclusions:
The test item has no skin sensitising potential in a skin sensitisation test with guinea pigs according to Draize.
Executive summary:

The skin sensitizing potential of the test item was assessed in an in a modified Draize skin sensitization test similar to OECD Guideline 406. Ten inbred albino guinea pigs of the Hartley strain, 350 g at the start of testing, were exposed to the test item by intradermal injection and topical application (non-occlusive). The test concentrations suitable for sensitization testing (Injection Challenge Concentration, ICC = 0.25%; Application Challenge Concentration, ACC = 10%) were derived from the results of a preliminary study. In the induction phase of the actual experiment, the test item was administered in 4 intradermal injections of 0.1 mL (at 4 sites overlying the 2 auxillary and 2 inguinal lymph nodes), each 2.5 times the derived ICC (i.e. 0.625%). After 14 days, the test animals were challenged intradermally with 0.1 mL of the test item at ICC and by topical application of 0.1 mL at ACC, spreading the test substance onto the shaved flank in a small circle. After 24 hours reactions were scored. On day 21 the induction procedure and on day 35 the challenge procedure was repeated, since no positive response was observable. The results were confirmed 7 days later by a third challenge (rechallenge), whereby a control was introduced comprising 4 previously untreated animals (similar weight and same sex to the test animals). The control animals were tretaed on the opposite flanks by intradermal injection and topical application of 0.1 mL at ICC and ACC, respectively. After 24 hours skin of the test group and control was examined for erythrema and oedema. The visualisation of skin reaction was achieved with a Philips colour-matching unit with 3 Philips 40 W Actinic Blue 05 fluorescent tubes and 3 Philips 40 W White 35 fluorescent tubes. The injection reaction was assessed by erythema and oedema of the 2 largest diameters; application reactions were scored on a 0 to +++ scale following the scoring system proposed by Draize. A positive reaction was considered when the total score of the test group was significantly greater than the average total score of the control group. Individual reactions were regarded as positive if  the score was + or greater and if there were no erythema found in the control. In result, there were no indications for a skin sensitising potential of the test item under the conditions chosen. Therefore, it can be concluded that the test item is not a skin sensitiser.

Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
experimental study
Adequacy of study:
weight of evidence
Study period:
1985
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
comparable to guideline study with acceptable restrictions
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 406 (Skin Sensitisation)
Version / remarks:
Open Epicutaneous Test (OET)
Deviations:
not applicable
Principles of method if other than guideline:
In vivo test with three consecutive phases: (1) pretesting phase in order to determine the primary irritating threshold concentration of test substance as baseline data, (2) induction phase of 3 weeks including daily non-occlusive applications of the test substance, (3) challenge phase to assess whether sensitisation has occured or not
- Parameters analysed / observed: Macroscopic signs of skin sensitisation in comparison to the control
GLP compliance:
no
Type of study:
open epicutaneous test
Justification for non-LLNA method:
Test performed 1985.
Species:
guinea pig
Strain:
not specified
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Not specified
- Females (if applicable) nulliparous and non-pregnant: Not specified
- Age at study initiation: Not specified
- Weight at study initiation: 300-450 g


ENVIRONMENTAL CONDITIONS
- Not specified
Route:
epicutaneous, open
Vehicle:
not specified
Concentration / amount:
100% (undiluted), 30%, 10%, 3%, 0.3%
Day(s)/duration:
21
Adequacy of induction:
highest concentration used causing mild-to-moderate skin irritation and well-tolerated systemically
No.:
#1
Route:
epicutaneous, open
Vehicle:
not specified
Concentration / amount:
0.025 mL/2 cm^2
A = min. irritating conc. (not further specified)
B = A : 3 max.nonlrritaling conc.
C = B : 3
D = C : 3
Day(s)/duration:
min. 48 h, max. 72 h
Adequacy of challenge:
other: highest non-irritant concentration, 3 lower concentrations and minimal irritating concentration to confirm biological activity
No. of animals per dose:
At least 6
Details on study design:
RANGE FINDING TESTS:
- Start:1 day before induction procedure
- Objective: To estimate the threshold irritating concentration of the test substance
- Volume: 0.025 mL of each test concentration
- Application: Simultaneously on one area measuring 2 cm^2
- Site: On the flank skin previously clipped and marked with circular stemps
- Reading: 24 h after application
- Criterion for the minimal irritating and maximal nonirritating concentration: all-or-none
- Minimal irritating concentration: Lowest concentration causing skin irritation
- Maximal non irritating concentration: Highest concentration not causing macroscopic skin reactions in any of the animals


MAIN STUDY
A. INDUCTION EXPOSURE
- No. of exposures: 20 - 21
- Exposure period: 21 - 28 days
- Test groups: 6
- Control group: 1
- Site: usually at the same sites (uncovered); application sites changed when very strong reactions noted
- Frequency of applications: a) daily for three weeks OR b) 5 times weekly in 4 weeks
- Concentrations: 100%, 30%, 10%, 3%,1% and 0.3 %

B. CHALLENGE EXPOSURE
- No. of exposures: 1
- Day(s) of challenge: 21 (or 28)
- Exposure period: 72 hours
- Test groups: 4
- Control group: 1
- Site: Contralateral flank, area: 2 cm^2
- Concentrations: A = min. irritating conc.; B = A : 3 max.non irritating conc.; C = B : 3; D = C : 3
- Evaluation (hr after challenge): 24, 48 and/or 72

Challenge controls:
- Number of test animals: 10 (previously untreated with test item and, if necessary, only treated with vehicle)
- Tested on days 21 and 35
- Application site: Contralateral flanks
- Concentrations: minimal irritating concentration (A) and three lower concentrations (B = A : 3; C 0 B : 3; D = C : 3)
Positive control substance(s):
no
Positive control results:
Positive control not performed.
Group:
negative control
Remarks on result:
no indication of skin sensitisation
Group:
positive control
Remarks on result:
not measured/tested
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
test chemical
Dose level:
0.0 mg/kg bw
No. with + reactions:
0
Total no. in group:
0
Clinical observations:
Not specified
Remarks on result:
no indication of skin sensitisation
Remarks:
dose level: B, C, D (not further specified)
Key result
Reading:
2nd reading
Hours after challenge:
48
Group:
test chemical
Dose level:
0.0 mg/kg bw
No. with + reactions:
0
Total no. in group:
0
Clinical observations:
Not specified
Remarks on result:
no indication of skin sensitisation
Remarks:
48 h and/or 72 h; dose level: B, C, D (not further specified)
Interpretation of results:
GHS criteria not met
Conclusions:
No sensitisation (-) was observed in an OET in guinea pigs.
Executive summary:

To assess the skin sensitizing potential of the test item, an in vivo study was carried out following an Open Epicutaneous Test Method similar to OECD Guideline 406. The test consisted of three consecutive phases: (1) A pretesting phase in order to determine the primary irritating threshold concentration of test substance as baseline data, (2) an induction phase of 3 weeks including daily (or on five days weekly in 4 weeks) non-occlusive epicutaneous applications of the test substance, (3) a challenge phase to assess whether sensitisation has occurred. All test solutions were applied with a pipette or a syringe. In the pretesting phase 0.025 mL of a 100, 30, 10, 3 and 1% test solution were administered non-occlusive on the flank skin which was previously clipped and marked with circular stamps. The reaction was read 24 h after application and a minimal irritating concentration as well as a maximal non irritating concentration (i.e. the highest concentration not causing macroscopic skin reactions in any of the animals) was derived. A negative or vehicle control with ten test animals ran in parallel. On day 1 the induction followed. Aliquots of 0.1 mL at concentrations of 100, 30, 10, 3,1 0.3 % were applied to an area of 8 cm2 daily for three weeks or 5 times weekly during four weeks. Usually, the application was performed at the same sites (uncovered), but when very strong reactions were noted, the application sites were changed. Reading was conducted 24 h after each application or at the end of each week. Simultaneously, a negative control with 10 previously untreated (or only treated with the vehicle) test animals was conducted. Like in the pre-test, the minimal irritating and maximal non-irritating concentration was determined by an all-or-none-criterion. The challenge phase began on day 21 or 28 respectively. An aliquot of 0.025 mL of the minimal irritating concentration (to exclude false results based on instability of the test material) and three lower concentrations (A = min. irritating conc.; B = A : 3; C = B : 3; D = C : 3) were administered non-occlusive to an area of 2 cm2 of the contralateral flank. Additionally a control of 10 previously untreated (or only treated with the vehicle) animals ran simultaneously, using the same concentration levels. Reading was performed 24, 48 and/or 72 hours after application. A positive response was considered at a concentration to which at least 1 of 6 animals showed signs of skin sensitization after challenging with nonirritating concentrations.

In result, no sensitisation was produced at a dose level of 6.0%. It was not specified if the concentration of 6.0% reported in the results was the minimal irritating concentration or 1 of the lower primary nonirritating concentration.

Endpoint:
skin sensitisation: in vivo (LLNA)
Type of information:
experimental study
Adequacy of study:
weight of evidence
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
data from handbook or collection of data
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)
Deviations:
not applicable
Principles of method if other than guideline:
- Name of Method: Murine local lymph node assay
- Endpoint and Indices: Lymphocyte proliferative responses provoked in draining lymph nodes after an epicutaneous application of the test item, quantified as 3H-TdR incorporation measured by ß-scintillation, expressed as disintegrations per minute (dpm). Stimulation Indices (SI) are determined as the increase in 3H-TdR relative to the vehicle control.
- Criteria for positive effect: A positive response is considered if a 3-fold increase in proliferation compared with the vehicle control occured at one or more test concentrations.

TREATMENT
1. Initial treatment: Treated once a day for three consecutive days on the dorsum of both ears with 25 µL of the test concentrations or with the vehicle alone
2. Five days later: Intravenous injection of in the tail vein of 250 µL phosphate buffered saline (PBS) containing containing 20 µCi of 3H-methylthymidine (3'H-TdR; specific activity 2.0 Ci/mmol)
3. After 5 hours: mice were euthanised, draining auricular lymph nodes were removed and pooled for each experimental group

CONTROLS
- Vehicle control: With acetone : olive oil , 4:1, without test item
- Positive control: Not performed


GLP compliance:
not specified
Type of study:
mouse local lymph node assay (LLNA)
Species:
mouse
Strain:
CBA/Ca
Sex:
female
Details on test animals and environmental conditions:
- Age: 6-12 weeks old
- Housed under standard conditions
- Methods approved by the U K Home Office and performed in compliance with the Animals (Scientific Procedures) Act 1986 (Home Office granted Project License)
Vehicle:
acetone/olive oil (4:1 v/v)
Concentration:
10%, 25% and 50% w/v
No. of animals per dose:
n = 4
Details on study design:
MAIN STUDY

ANIMAL ASSIGNMENT AND TREATMENT
- Name of test method: Murine local lymph node assay
- Criteria used to consider a positive response: if a 3-fold increase in proliferation compared with the vehicle control occurs at least one test concentration

TREATMENT PREPARATION AND ADMINISTRATION:
1. Initial treatment: Treated once a day for three consecutive days on the dorsum of both ears with 25 µL of the test concentrations or with the vehicle alone
2. Five days later: Intravenous injection of in the tail vein of 250 µL phosphate buffered saline (PBS) containing containing 20 µCi of 3H-methylthymidine (3'H-TdR; specific activity 2.0 Ci/mmol)
3. After 5 hours: mice were euthanised, draining auricular lymph nodes were removed and pooled for each experimental group

Positive control substance(s):
not specified
Statistics:
N/A
Positive control results:
N/A
Key result
Parameter:
SI
Value:
1.3
Variability:
Not specified
Test group / Remarks:
Concentration: 10% w/v
Key result
Parameter:
SI
Value:
1
Variability:
Not specified
Test group / Remarks:
Concentration: 25% w/v
Key result
Parameter:
SI
Value:
1
Variability:
Not specified
Test group / Remarks:
Concemtration: 50% w/v
Cellular proliferation data / Observations:
N/A

Table: Results of the LLNA test.

Vehicle and
Concentrations [% w/v]		 dpm/node Stimulation index Sensitization classification
 by LLNA		 Human sensitisation classification
AOO 392 1
10 490 1.3 - -
25 404 1.0
50 395 1.0

AOO = acetone : oilive oil, 4 : 1

dpm = disintegrations per minute

Interpretation of results:
GHS criteria not met
Conclusions:
The test item has no skin sensitising potential in a LLNA test
Executive summary:

To assess the skin sensitizing potential of the test item a Murine Local Lymph Node Assay similar to OECD Guideline 429 was performed (2000). Female CBA/J strain mice or female CBA/Ca strain mice, age 6-12 weeks old, were used in these studies. Test chemicals were chosen for this study based on two criteria: (i) the potential of the chemical to induce contact allergy in humans was well established; and (ii) the chemical had not been previously tested in the local lymph node assay (LLNA). Groups of mice (n = 4 or 5) were treated once per day for 3 consecutive days on the dorsum of both ears with 25 ul of 1 of 3 concentrations of test material or with vehicle along. 5 days after the initial treatment, all mice were injected intravenously, via the tail vein, with 250 ul of phosphate buffered saline (PBS) containing 20 uCi of [3H]-methylthymidine. Five hours later, the mice were euthanized and the draining auricular lymph nodes were removed and pooled for each individual mouse or for each experimental group. Single cell suspensions were prepared by gentle mechanical disaggregating through either nylon mesh (100 um pore size) or 200-mesh stainless steel gauze. Cell suspensions were washed 2X with an excess of PBS and precipitated with 5% trichloroacetic acid (TCA) and left at 4 C overnight. The samples were then centrifuged, resuspended in 1 ml 5% TCA and transferred to 10 ml of scintillation cocktail. [3H]TdR incorporation was measured by B-scintillation counting and expressed as disintegrations per min (dpm) per individual mouse or as dpm per node for each experimental group. Stimulation indices (SI) for each experimental group were determined as the increase in [3H]TdR incorporation relative to the concurrent vehicle treated control group. A material is considered to be positive in the LLNA if 3-fold or greater increase in proliferation, compared with the concurrent vehicle control, is obtained at 1 or more test concentrations. In result the SI for the 10%-level was 1.3 and 1.0 both for the 25%-level 1.0 and the highest concentration of 50% w/v. It can be concluded that the test item has no skin sensitizing potential, since the SI for all tested concentrations was lower than 3.0 and therefore no positive response is considered. In result the SI for the 10%-level was 1.3 and 1.0 both for the 25%-level 1.0 and the highest concentration of 50% w/v. It can be concluded that the test item has no skin sensitizing potential, since the SI for all tested concentrations was lower than 3.0 and therefore no positive response is considered.

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

Skin sensitisation

In vivo tests

Murine Local Lymph Node Assay

To assess the skin sensitizing potential of the test item a Murine Local Lymph Node Assay similar to OECD Guideline 429 was performed (2000). For this study 6-12 weeks old mice of the CBA/Ca strain were tested (4 mice per treatment group). The test item was administered at three concentrations of 10%, 25% and 50% w/v with acetone : olive oil (4:1) as vehicle. A vehicle control was performed in parallel. The test animals were treated once a day for three consecutive days on the dorsum of both ears with 25 µL of the test concentrations or with the vehicle alone. Five days later 250 µL phosphate buffered saline (PBS) containing 20 µCi of 3H-methylthymidine (3'H-TdR; specific activity  2.0 Ci/mmol) were injected intravenously in the tail vein. Five hours after application all mice were euthanized. Afterwards, draining auricular lymph nodes were removed and pooled for each experimental group. Lymphocyte proliferative responses were quantified as3H-TdR incorporation measured by ß-scintillation and expressed as disintegrations per minute (dpm). Stimulation Indices (SI) were determined as the increase in3H-TdR relative to the vehicle control (SI = 1). A positive response was considered if a 3-fold increase in proliferation compared with the vehicle control occurred at one or more test concentrations. In result the SI for the 10%-level was 1.3 and 1.0 both for the 25%-level 1.0 and the highest concentration of 50% w/v. It can be concluded that the test item has no skin sensitizing potential, since the SI for all tested concentrations was lower than 3.0 and therefore no positive response is considered.

Modified Draize Test

Sensitization potential was measured in a guinea pig sensitization study using a modified Draize procedure (1978). Ten male and female inbred Hartley strain albino guinea pigs/group with an average weight of 350 grams were shaved on both flanks. A 0.1 ml aliquot of test material, at 2.5 times the ICC, was injected intradermally at 4 sites which overlap the 2 auxiliary and 2 inguinal lymph nodes. The guinea pigs were challenged 14 days later by an intradermal injection of 0.1 ml test material into one flank and a topical open application of test material on the other flank at the respective injection challenge concentration (ICC) and application challenge concentration (ACC). Reactions were scored 24 hours after challenge treatments. A second challenge was carried out 7 days later. If no sensitization reactions were observed, the test was repeated, but this time a confirmatory challenge with controls was included. At each challenge with controls, 4 previously untreated animals of the same sex and similar weight to the test animals were treated intradermally and topically on opposite flanks with 0.1 ml aliquots of test substance at the ICC and ACC respectively. Reactions were examined under a Philips colour-matching unit. Each injection was given a total score based on size (2 largest diameters), erythema and oedema. Individual reactions were considered positive when their total score was significantly greater than the average total score for control reactions. Application reactions were scored on a 0 to +++ scale and individual reactions were considered positive if they were + or greater and there were no erythema reactions in controls. At an ACC-dose of 10% and an ICC-dose of 0.25% no signs of skin sensitisation were observeable.

Open Epicutaneous Test (OET)

A guinea pig Open Epicutaneous Test (OET) was conducted (1985) on groups of 6 - 8 male and female guinea pigs weighting 300 - 450 grams. Daily applications were made for 3 weeks to a clipped 8 cm2 area on the flank of each guinea pig. The test sites were not covered and the reactions were read 24 hours after each application. A total of 21 applications of 0.1 ml test material in an unspecified vehicle were made for 21 days. The 10 controls were either left untreated or treated with 0.1 ml of the vehicle for 21 days. At the challenge phase, both the test and control animals were treated on days 21 and 35 on the contralateral flank with the test material at the minimal irritating concentration and some lower primary non-irritating concentrations.

Annother open epicutaneous test was conducted in guinea pigs (1979). Induction consisted of 21 daily open applications to the shaved flank of 6-8 guinea pigs per group. One to six experimental and one control group was used. Open challenge applications were made on days 21 and 35. Reactions were read at 24, 48 and 72 hours. No further details were provided. At a dose level of 6% no signs of skin sensitisation were noted.

Conclusion: 

Skin sensitizing studies on guinea pigs and a human patch test from Kligmann (1973; please refer to section 7.10.4) showed no sensitizing potential under the test conditions chosen. Therefore, the substance is considered to be not classified according to Regulation (EC) No 1272/2008.

Respiratory sensitisation

Endpoint conclusion
Endpoint conclusion:
no study available

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 on skin sensitisation, the test item is not considered to be classified for skin sensitisation according to Regulation (EC) No 1272/2008 (CLP), as amended for the fifteenth time in Regulation (EU) 2020/1182.