Alcohols, C12-14, ethoxylated

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (non-LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

16 Sep - 27 Oct 1996

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1996

Report date:

1997

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 406 (Skin Sensitisation)

Version / remarks:

adopted in 1992

Deviations:

yes

Remarks:

10 test and 5 control animals per group were used instead of 20 test and 10 control animals. Since there is a second test on skin sensitiation in guinea pigs available, no additional testing was performed in the present study.

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.6 (Skin Sensitisation)

GLP compliance:

yes (incl. QA statement)

Remarks:

The Department of Health of the Government of the United Kingdom

Type of study:

guinea pig maximisation test

Justification for non-LLNA method:

Test was done before LLNA as first-choice method for in-vivo testing was set into force.

Test material

Test material information

Constituent 1

Reference substance name:

Alcohols, C12-14, ethoxylated

EC Number:

500-213-3

EC Name:

Alcohols, C12-14, ethoxylated

Cas Number:

68439-50-9

Molecular formula:

n.a.

IUPAC Name:

Alcohols, C12-14, ethoxylated, < 2.5 EO

In vivo test system

Test animals

Species:

guinea pig

Strain:

Dunkin-Hartley

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: David Hall Limited, Staffordshire, United Kingdom
- Age at study initiation: 12 weeks
- Weight at study initiation: 329 - 427g
- Housing: singly or in pairs in solid-floor polypropylene cages furnished with woodflakes
- Diet: Guinea Pig FD1 Diet (Special Diet Services Limited, Witham, Essex, United Kingdom), ad libitum
- Water: tap water, ad libitum
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18-22
- Humidity (%): 47-68
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 12/12

Study design: in vivo (non-LLNA)

Induction

Route:

intradermal and epicutaneous

Vehicle:

arachis oil

Concentration / amount:

Intradermal Induction: 1%
Topical Induction: 100%

Day(s)/duration:

epicutaneous induction: 48 h

Adequacy of induction:

other: induction doses based on dose-range finding test using concentrations of 1 and 5% for dermal injection and 25, 50, 75 and 100% for epicutaneous induction

Challenge

No.:

#1

Route:

epicutaneous, occlusive

Vehicle:

arachis oil

Concentration / amount:

Topical Challenge: 25 and 50%

Day(s)/duration:

24 h

Adequacy of challenge:

other: dose levels for topical challenge were selected based on the results of a dose-range finding test; the highest concentration was the highest non-irritant concentration

No. of animals per dose:

10 (in test group), 5 (control)

Details on study design:

RANGE FINDING TESTS:
Two guinea pigs were dermally injected with test item concentrations of 1 and 5%. The animal induced with the 1% formulation showed erythema of grade 3 at the 24, 48 and 72 h reading and erythema of grade 2 at the 7 day reading. The animal induced with the 5% formulation showed erythema of grade 3-4 and green necrosis 24 h after induction and erythema of grade 4 with green necrosis findings and eschar formation at the 48 h, 72 h and 7 day reading. Thus, the 1% formulation was chosen for intradermal induction.
In addition, two animals were topically treated with 25, 50, 75 and 100% concentrations for 48 h. One hour after induction, both animals showed erythema of grade 2-3 at all concentration levels. Edema of grade 2 was observed for both animals at 100% and for one animal at 75 and 50%. Edema of grade 1 was noted in one animal at 25% and at 50 and 75% in the second animal. At the 24 h reading, erythema persisted in both animals at the 75% (score 1) and 100% formulation (score 2) and in one animal at the 50% formulation (score 1). One of the animals shiwed edema formation (grade 1) at 100 and 50%, whereas no edema was observed in the second animal.
At the 48 h reading, both animals showed erythema of grade 2 at the 100% formulation test site and erythema of grade 1 at the 75% formulationt test site. In one of the animals, edema was observed (grade 1) at the 100% formulation test site.
Two additional animals were used for a sighting test on topical challenge. One hour after challenge, erythema of grade 2 was noted in both animals at 100 and 75% and erythema of grade 1 was observed at 50 and 25% in both animals. Edema of gradde 1 was noted at 100% in both animals and at 50% in one of the guinea pigs. At the 24 h reading, both animals showed erythema of grade 1 when treated at 50 and 100%. Erythma of score 1 at challenge wirh 100% persisted in one of the animals up to the 48 h time point. At the 24 and 48 h reading, no finding of edema was observed in any animal.

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): 0.1 mL each
Injection 1: 1:1 mixture complete Freund's adjuvant (FCA) and distilled water
Injection 2: 1% test substance in arachis oil BP
Injection 3: 1% test substance in a 1:1 mixture of complete Freund's adjuvant (FCA) and distilled water
Epicutaneous: filter paper patch saturated with undiluted test material
- Control group:
Intradermal (3 pairs of injections):
Injection 1: 1:1 mixture complete Freund's adjuvant (FCA) and distilled water
Injection 2: 100% arachis oil BP
Injection 3: a 50% formulation of arachis oil BP in a 1:1 mixture of complete Freund's adjuvant (FCA) and distilled water
Epicutaneous: filter paper patch saturated with arachis oil BP
- Site: shoulder region
- Frequency of applications: topical induction was performed one week after the intradermal induction
- Duration: 7 days
- Concentrations: intradermal 1%, epicutaneous 100%

B. CHALLENGE EXPOSURE
- No. of exposures: 1
- Day(s) of challenge: 3 weeks after intradermal induction (study Day 21)
- Exposure period: 24 h
- Test groups: test substance and vehicle
- Control group: test substance and vehicle
- Site: left flank (test item) and right flank (vehicle)
- Concentrations: 25 and 50%
- Evaluation (hr after challenge): 48 and 72 h (24 and 48 h after dressing removal)

Challenge controls:

The control group is actually a challenge control

Positive control substance(s):

yes

Remarks:

Historical data on ethyl-4-aminobenzoate, 2,4-dinitrochlorobenzene, neomycin sulphate and 2-mercaptobenzothiazole were provided.

Results and discussion

Positive control results:

Historical control data on several positive control chemicals were provided. Challenge with 2-mercaptobenzothiazole (intrademally induced with 10% in arachis oil, topically induced with 50% in acetone and topically challenged with 25 and 50% in acetone : PEG 400 in a 70:30 ratio), exhibited dermal alterations in 70% of the test animals, thus meeting the reliability criteria for the GPMT test (≥ 15% positive reactions).

In vivo (non-LLNA)

Results

Results 1

Key result

Reading:

1st reading

Hours after challenge:

24

Group:

test chemical

Dose level:

intradermal induction: 1%; epicutaneous induction: 100%; epicutaneous challenge: 25%

No. with + reactions:

0

Total no. in group:

10

Results 2

Key result

Reading:

2nd reading

Hours after challenge:

48

Group:

test chemical

Dose level:

intradermal induction: 1%; epicutaneous induction: 100%; epicutaneous challenge: 25%

No. with + reactions:

0

Total no. in group:

10

Results 3

Reading:

1st reading

Hours after challenge:

24

Group:

negative control

Dose level:

challenge: 25%

No. with + reactions:

0

Total no. in group:

5

Results 4

Reading:

2nd reading

Hours after challenge:

48

Group:

negative control

Dose level:

challenge: 25%

No. with + reactions:

0

Total no. in group:

5

Results 5

Key result

Reading:

1st reading

Hours after challenge:

24

Group:

test chemical

Dose level:

intradermal induction: 1%; epicutaneous induction: 100%; epicutaneous challenge: 50%

No. with + reactions:

0

Total no. in group:

10

Results 6

Key result

Reading:

2nd reading

Hours after challenge:

48

Group:

test chemical

Dose level:

intradermal induction: 1%; epicutaneous induction: 100%; epicutaneous challenge: 50%

No. with + reactions:

0

Total no. in group:

10

Results 7

Reading:

1st reading

Hours after challenge:

24

Group:

negative control

Dose level:

challenge: 50%

No. with + reactions:

0

Total no. in group:

5

Results 8

Reading:

2nd reading

Hours after challenge:

48

Group:

negative control

Dose level:

challenge: 50%

No. with + reactions:

0

Total no. in group:

5

Results 9

Group:

positive control

Dose level:

intradermal induction: 10%; epicutaneous induction: 50%; epicutaneous challenge: 25 and 50%

Remarks on result:

other: Historical positive control data provided in study report

Any other information on results incl. tables

Local skin reactions:

After intradermal induction, well-defined or moderate to severe erythema was noted at the induction sites of all test group animals at the 24- and 48-h observation. Very slight erythema was noted at the intradermal induction sites of 4 control animals at the 24 and 48 h reading.

After topical induction, very slight to moderate to severe erythema with or without very slight edema was noted at the induction sites of all test group animals at the 1-h observation. Very slight to well-defined erythema with or without very slight to slight edema was noted at the induction sites of seven test group animals at the 24-h observation. Dried blood was noted at the induction sites of six test group animals at the 24 -h observation. Skin reactions prevented the accurate evaluation of erythema and edema at the induction sites of two test group animals at the 24 -h observation. No skin reactions were noted at the treatment sites of control group animals at the 1- and 24-h observations.

After topical challenge, no skin reactions were observed in any control or test item-treated animal at the 24 and 48 -h observation.

Table 1: Skin sensitisation results

Animals Number	skin Reactions (Hours After Removal of Dressing)
	24 h						48 h
	50%			25%			50%			25%
	Er	Oe	Other	Er	Oe	Other	Er	Oe	Other	Er	Oe	Other
1	0	0	-	0	0	-	0	0	-	0	0	-
2	0	0	-	0	0	-	0	0	-	0	0	-
3	0	0	-	0	0	-	0	0	-	0	0	-
4	0	0	-	0	0	-	0	0	-	0	0	-
5	0	0	-	0	0	-	0	0	-	0	0	-
6	0	0	-	0	0	-	0	0	-	0	0	-
7	0	0	-	0	0	-	0	0	-	0	0	-
8	0	0	-	0	0	-	0	0	-	0	0	-
9	0	0	-	0	0	-	0	0	-	0	0	-
10	0	0	-	0	0	-	0	0	-	0	0	-

Er=erythema

Oe=oedema

- = no other reactions noted

Er=erythema

Oe=oedema

- = no other reactions noted

Systemic toxicity: Not reported.

Body weight: All animals gained weight as expected.

Applicant's summary and conclusion

Interpretation of results:

other: CLP/EU GHS criteria not met, no classification required according to Regulation (EC) No. 1272/2008.

Executive summary:

A study was performed to assess the contact sensitisation potencial of the test material in the albino guinea pig. Ten test and five control animals were used for the main study. Based on the results of sighting test, the concentration of the test material for the induction and challenge phases were selected as follows:

Intradermal Induction: 1% w/v in arachis oil BP

Topical Induction: undiluted as supplied

Topical Challenge: 50% and 25% v/v in arachis BP

The test material produced a 0% (0/10) sensitisation rate and was classified as a NON-SENSITISER to guinea pig skin. The test material did not meet the criteria for classification as a sensitiser according to EU labelling regulations. No risk phrase is required.