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Description of key information

In the key study, the skin sensitisation properties of 2-ethylhexyl benzoate were tested in a GPMT according to OECD guideline 406 in compliance with GLP (Coleman, 2000).

In a preliminary range finding test, the suitable concentrations were evaluated for the intradermal injection and the patch testing. In the main study, Dunkin-Hartley guinea pigs were induced with a single intradermal injection of the test substance at 7.5% in Alembicol D and an epicutaneous occlusive application of the undiluted test substance at the flank 7 days later. Epicutaneous challenge exposure was conducted 20 days after the first induction for 24 h under occlusive conditions at 37.5 and 75% of the test substance applied on the flanks.

All test and control animals except one showed no skin reactions after 24 and 48 h. 1/10 test group animals showed a slight erythema at the 75% application site after 24 h. The positive control group was treated with 10% hexyl cinnamic aldehyde in Alembicol D and showed the expected results, thus confirming the sensitivity and reliability of the experiment. 

Under the experimental conditions described, it was concluded, that no evidence of skin sensitisation properties were seen after treatment with the 2-ethylhexyl benzoate.

Furthermore, in a short abstract, the skin sensitisation properties of 2-ethylhexyl benzoate are described in an assay equivalent to OECD guideline 429 (Johnson, 2001).

In the local lymph node assay, the radiolabelled thymidine incorporation method was used. Concentrations of 3, 10 and 30% (w/v) of the test material in acetone in olive oil were tested in mice.

The resulting test/control ratios from the treatment groups were below the value which is recognized for a positive response (1.07, 1.00 and 2.41).

In summary, under the conditions described, no evidence of skin sensitisation properties was seen after treatment with 2-ethylhexyl benzoate.

Key value for chemical safety assessment

Skin sensitisation

Link to relevant study records

Referenceopen allclose all

Endpoint:
skin sensitisation: in vivo (LLNA)
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
4 (not assignable)
Rationale for reliability incl. deficiencies:
other: Only short abstract available. Lack of details on test material and method.
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)
Deviations:
yes
Remarks:
lack of details on test material and method
GLP compliance:
yes
Remarks:
study protocol, experimental phases and report were not subject of Quality Assurance audit
Type of study:
mouse local lymph node assay (LLNA)
Species:
mouse
Strain:
not specified
Sex:
not specified
Vehicle:
other: acetone in olive oil
Concentration:
3, 10 and 30% (w/v)
No. of animals per dose:
no data
Details on study design:
MAIN STUDY
ANIMAL ASSIGNMENT AND TREATMENT
- Name of test method: radiolabelled thymidine incorporation
Key result
Parameter:
other: Test / Control Ratio (CPM)
Value:
< 2.5

Table 1. Results of local lymph node assay.

Concentration of

test material

CPM/lymph node (x10-2)

Test/control ratio

Vehicle

1.67

N/A

3% (w/v)

1.78

1.07

10% (w/v)

1.67

1.00

30% (w/v)

4.02

2.41

CPM - counts per minute

Interpretation of results:
GHS criteria not met
Conclusions:
According to the authors, the test material is considered unlikely to be a skin sensitiser under the conditions of the test.
Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
experimental study
Adequacy of study:
key study
Study period:
24 Jan - 25 Feb 2000
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP-Guideline study.
Qualifier:
according to
Guideline:
OECD Guideline 406 (Skin Sensitisation)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.6 (Skin Sensitisation)
Deviations:
no
Qualifier:
according to
Guideline:
EPA OPPTS 870.2600 (Skin Sensitisation)
Deviations:
no
GLP compliance:
yes
Type of study:
guinea pig maximisation test
Justification for non-LLNA method:
Vertberate study predates requirement to test in-vitro before vertebrate studies
Species:
guinea pig
Strain:
Dunkin-Hartley
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: D. Hall (Newchurch, Staffs, UK)
- Age at study initiation: 4-7 weeks
- Weight at study initiation: 362-435 g
- Housing: the animals were housed in groups of five in suspended metal cages with wire mesh floors.
- Diet: vitamin C enriched guinea-pig diet (Harlan Teklad 9600 FD2 SQC), ad libitum. In addition, hay was given thrice weekly.
- Water: drinking water, ad libitum
- Acclimation period: 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19-20
- Humidity (%): 38-58
- Photoperiod (hrs dark / hrs light): 12/12

Route:
intradermal and epicutaneous
Vehicle:
other: Alembicol D
Concentration / amount:
Preliminary study:
Intradermal: 0.1-10%; epicutaneous: 25-100%
Main study:
Induction:
Intradermal: 7.5%; epicutaneous: 100%
Challenge:
37.5 and 75%
Route:
epicutaneous, occlusive
Vehicle:
other: Alembicol D
Concentration / amount:
Preliminary study:
Intradermal: 0.1-10%; epicutaneous: 25-100%
Main study:
Induction:
Intradermal: 7.5%; epicutaneous: 100%
Challenge:
37.5 and 75%
No. of animals per dose:
Preliminary study
6
Main study
5 (controls), 10 (in test group)
Details on study design:
MAIN STUDY
A. INDUCTION EXPOSURE
- No. of exposures: 2 (intradermal and epicutaneous, respectively)
- Exposure period: single injection (intradermal) and 48 h (epicutaneous)

- Test groups:
Intradermal (3 pairs of injections):
Injection 1: a 1:1 mixture (v/v) FCA/water
Injection 2: test substance in Alembicol D
Injection 3: test substance in a 1:1 mixture (v/v) FCA/water
Epicutaneous: test substance

- Control group:
Intradermal (3 pairs of injections):
Injection 1: a 1:1 mixture (v/v) FCA/water
Injection 2: Alembicol D
Injection 3: Alembicol D in a 1:1 mixture (v/v) FCA/water
Epicutaneous: Alembicol D

- Site: flank
- Frequency of applications: every 7 days
- Duration: Day 0-8
- Concentrations: intradermal 7.5%, epicutaneous 100%

B. CHALLENGE EXPOSURE
- No. of exposures: 1
- Day(s) of challenge: 20
- Exposure period: 24 h
- Test groups: test substance
- Control group: test substance
- Site: left flank
- Concentrations: 37.5 and 75% (v/v)
- Evaluation (hr after challenge): 48 and 72 h

OTHER:
The preliminary investigations indicated that the neat test substance applied topically did not produce skin irritation. Therefore, six days after the injections, the injection site was clipped and shaved free of hair and the site was pre-treated by gentle rubbing with 0.5 mL per site of 10% (w/w) sodium lauryl sulphate in petrolatum.
Challenge controls:
The control group is actually a challenge control.
Positive control substance(s):
yes
Remarks:
hexyl cinnamic aldehyde (induction: 10% intradermal; epicutaneous 100%; challenge: 100%)
Positive control results:
The positive control substance (10% hexyl cinnamic aldehyde in Alembicol D (intradermal), 50 and 100% (challenge)) induced positive reactions in 9/10 animals (90%), thus confirming the sensitivity and reliability of the experimental technique.
Reading:
1st reading
Hours after challenge:
24
Group:
negative control
Dose level:
0% (induction); 37.5 and 75% (challenge)
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: 0% (induction); 37.5 and 75% (challenge). No with. + reactions: 0.0. Total no. in groups: 5.0.
Reading:
1st reading
Hours after challenge:
24
Group:
test group
Dose level:
7.5 and 100% (induction: intradermal and epicutaneous); 37.5 and 75% (challenge)
No. with + reactions:
1
Total no. in group:
10
Clinical observations:
slight erythema at application site (75% of the test material) in 1/10 animals
Remarks on result:
other: see Remark
Remarks:
Reading: 1st reading. . Hours after challenge: 24.0. Group: test group. Dose level: 7.5 and 100% (induction: intradermal and epicutaneous); 37.5 and 75% (challenge). No with. + reactions: 1.0. Total no. in groups: 10.0. Clinical observations: slight erythema at application site (75% of the test material) in 1/10 animals.
Reading:
2nd reading
Hours after challenge:
48
Group:
negative control
Dose level:
0% (induction); 37.5 and 75% (challenge)
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: 0% (induction); 37.5 and 75% (challenge). No with. + reactions: 0.0. Total no. in groups: 5.0.
Reading:
2nd reading
Hours after challenge:
48
Group:
test group
Dose level:
7.5 and 100% (induction: intradermal and epicutaneous); 37.5 and 75% (challenge)
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: 7.5 and 100% (induction: intradermal and epicutaneous); 37.5 and 75% (challenge). No with. + reactions: 0.0. Total no. in groups: 10.0.

No signs of ill health or toxicity were observed.

After intradermal injections with FCA necrosis was recorded in test and control animals. Slight irritation was seen in test animals receiving intradermal 7.5% of the test substance in Alembicol D and in control animals.

After topical application, slight erythema was observed in 5/10 animals and well defined erythema in 4/10 animals; no erythema was seen in the control animals. After challenge, slight erythema was noted for one test animal at the 24 h reading only. The reaction was considered to be a background irritation.

Interpretation of results:
GHS criteria not met
Conclusions:
CLP: not classified
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (not sensitising)
Additional information:

Justification for selection of skin sensitisation endpoint:
The key study is GLP compliant and of high quality (Klimisch score = 1).

Respiratory sensitisation

Endpoint conclusion
Endpoint conclusion:
no study available
Additional information:

This information is not available.

Justification for classification or non-classification

The available data on the skin sensitising potential of 2-ethylhexyl benzoate do not meet the criteria for classification according to Regulation (EC) 1272/2008 or Directive 67/548/EEC, and are therefore conclusive but not sufficient for classification.