Registration Dossier

Diss Factsheets

Administrative data

Description of key information

The test substance has no skin sensitising effects.

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
Study period:
From May 13, 2002 to May 21, 2002
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)
Deviations:
yes
Remarks:
male mice instead of female mice were used; groupe housing instead of single housing; no body weight measurement
GLP compliance:
yes
Type of study:
mouse local lymph node assay (LLNA)
Species:
mouse
Strain:
CBA/Ca
Sex:
male
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Harlan, Interfauna UK Limited, Blackthorne, Bicester, Oxon, UK
- Sex: male
- Age at study initiation: young adults
- Weight at study initiation:
- Housing: 4/cage
- Diet (e.g. ad libitum): RM1 ad libitum
- Water (e.g. ad libitum): tap water ad libitum
- Acclimation period: at least 5 d
- Indication of any skin lesions:

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22+/-3
- Humidity (%): 30-70
- Air changes (per hr): minimum 15
- Photoperiod (hrs dark / hrs light): 12/12
- IN-LIFE DATES: The main study was initiated on 13 May 2002. The experimental phase started on 14 May 2002 and was completed on 21 May 2002.
For the positive control study, the experimental phase started on 18 September 2001 and was completed on 25 September 2001.
Vehicle:
other: acetone
Concentration:
3%, 10% and 50% w/v
No. of animals per dose:
groups of 4 males
Details on study design:
Groups of four male mice were used for this study. Approximately 25 µL of a 3%, 10% or 30% w/v preparation of the test substance in acetone was applied, using a variable volume micro-pipette, to the dorsal surface of each ear. A vehicle control group was similarly treated using acetone alone. The procedure was repeated daily for 3 consecutive days.
Three days after the third application, all animals were injected, via the tail, with approximately 250 µL of PBS containing about 20 µCi of a 2.0Ci/mmol specific activity 3H-methyl thymidine (for beta-scintillation counting using a Packard Tri-Carb 2500TR Liquid Scintillation Counter). After 5 d, the animals were sacrificed. The draining auricular lymph nodes were removed from each animal and, together with the nodes from the other animals in the group, were placed in a container of PBS.

The results are expressed as a counts per minute (cpm) value per lymph node for each group. The stimulation index for each test group is then calculated by dividing the cpm value per lymph node by the equivalent value for the control (vehicle only) group.
The criterion for a positive response is that one or more concentrations of the test substance should elicit a 3-fold or greater increase in isotope incorporation relative to the vehicle control group. The assay is able to identify those materials that elicit responses in standard guinea pig tests for skin sensitisation (Kimber et al 1994). Consequently, a test substance which does not fulfil the above criterion is designated as unlikely to be a sensitiser.


Positive control substance(s):
hexyl cinnamic aldehyde (CAS No 101-86-0)
Positive control results:
The application of hexylcinnamaldehyde at concentrations of 1, 3 and 10% w/v in acetone resulted in a greater than 3-fold increase in isotope incorporation at all three concentrations.

The validity of the protocol was confirmed.
Key result
Parameter:
SI
Value:
0.83
Test group / Remarks:
3% of test substance
Key result
Parameter:
SI
Value:
1.19
Test group / Remarks:
10% of test substance
Key result
Parameter:
SI
Value:
1.06
Test group / Remarks:
30% of test substance
Cellular proliferation data / Observations:
The application of the test substance at concentrations of 3%, 10% and 30% w/v in acetone resulted in an increase in isotope incorporation which was less than 3-fold at all three concentrations.

Concentration of
test substance
(% w/v)

Number of
lymph nodes
assayed

Counts per
minute

(cpm)

cpm per
lymph node
(x10-2)

Test control
ratio

0 (vehicle only)

8

1092

1.37

N/A

3

8

913

1.14

0.83

10

8

1303

1.63

1.19

30

8

1160

1.45

1.06

Interpretation of results:
GHS criteria not met
Conclusions:
Under the study conditions, the test substance was considered to be not sensitizing to mouse skin (LLNA).
Executive summary:

A study was conducted to determine the skin sensitisation potential of the test substance according to OECD Guideline 429 (local lymph node assay), in compliance with GLP. Male CBA/Ca mice were exposed to the test substance at concentrations of 0, 3, 10 and 30% in acetone. Isotope (3H-methyl thymidine) incorporation was measured in lymph nodes. The negative (vehicle alone) and positive (hexylcinnamaldehyde at concentrations of 1, 3, and 10% w/v in acetone) controls were valid. The isotope incorporation was increased by less than a threefold at all concentrations of the test substance. Under the study conditions, the test substance was considered to be not sensitizing to mouse skin (Johnson, 2002).

Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
experimental study
Adequacy of study:
key study
Study period:
December 1978
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
comparable to guideline study with acceptable restrictions
Reason / purpose for cross-reference:
reference to same study
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 406 (Skin Sensitisation)
GLP compliance:
no
Remarks:
Study pre-dates GLP
Type of study:
not specified
Justification for non-LLNA method:
Study available is over 12 years old.
Species:
guinea pig
Strain:
Hartley
Sex:
female
Details on test animals and environmental conditions:
Ten young female Hartley-Dunkin albino guinea pigs obtained from Redfern Animal Breeders, Jasons Farm, Old Hay, Brenchley, Kent were used for this study. They were conditioned to the laboratory environment for not less than 6 days and on the day before initial treatment they were weighed. At this time the body weights of the guinea pigs were within the range 300 - 430 g. Three animals were marginally over the body weight range required by the protocol.

Diet
The animals were allowed free access to amins water and food (Guinea pig Diet-Standard supplemented with Vit. C., BP Nutrition (U.K.) Ltd., Stepfield, Witham, Essex).

Environment
All animals were housed in a single air-conditioned room maintained at a temperature of 22 + 3°C, relative humidity 50 + 10% and exposed to natural lighting conditions. They were caged in groups of 2 in grid floor polypropylene boxes.
Route:
epicutaneous, open
Vehicle:
N,N-dimethylformamide
Concentration / amount:
0.1 ml of a 10% w/v solution
Day(s)/duration:
3 days
Adequacy of induction:
not specified
No.:
#1
Route:
epicutaneous, open
Vehicle:
N,N-dimethylformamide
Concentration / amount:
0.2 ml
10% w/v, 1% w/v and 0.1% w/v
Day(s)/duration:
1 day
No. of animals per dose:
6 animals
Details on study design:
The test article (0.1 ml of a 10% w/v solution in dimethylformamjde) was applied daily by means of a glass syringe to the outer surface of the ears of 6 guinea pigs (animal numbers 1 - 6) for three days
(days 1, 2 and 3). On Day 8, 0.2 ml of the challenge solutions (10% w/v, 1% w/v and 0.1% w/v in dimethylformamide) was applied topically to 1 cm diameter circular areas on the clipped flanks of each of the same 6 animals. Solutions of test article were also applied in the same way on Day 8 to the clipped flanks of control animals (animal numbers 7 - 10) which had no previous treatment on the ears. The-applications were made on both flanks of all 10 guinea pigs, with each concentration being applied to each flank. The highest concentration was applied closest to the posterior end of the animal while the lowest concentration was applied nearest the anterior end.
The erythema produced on each site was assessed 24 hours later (Day 9) and graded on a 6 point scale.
Challenge controls:
The highest concentration was applied closest to the posterior end of the animal while the lowest concentration was applied nearest the anterior end.
Positive control substance(s):
not specified
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
test chemical
Dose level:
10% w/v
No. with + reactions:
0
Total no. in group:
6
Clinical observations:
None specified
Remarks on result:
no indication of skin sensitisation
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
test chemical
Dose level:
1% w/v
No. with + reactions:
0
Total no. in group:
6
Clinical observations:
None specified
Remarks on result:
no indication of skin sensitisation
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
test chemical
Dose level:
0.1% w/v
No. with + reactions:
0
Total no. in group:
6
Clinical observations:
None specified
Remarks on result:
no indication of skin sensitisation
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
negative control
Dose level:
10% w/v
No. with + reactions:
0
Total no. in group:
4
Clinical observations:
None specified
Remarks on result:
no indication of skin sensitisation
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
negative control
Dose level:
1.0% w/v
No. with + reactions:
0
Total no. in group:
4
Clinical observations:
None specified
Remarks on result:
no indication of skin sensitisation
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
negative control
Dose level:
0.1% w/v
No. with + reactions:
0
Total no. in group:
4
Clinical observations:
None specified
Remarks on result:
no indication of skin sensitisation

Skin reactions at challenge phase

Test article: Disperse Brown 19

Vehicle: dimethylformamide

Concentration: 10% w/v, 1% w/v and 0.1% w/v

Animal number and sex

Weight

(g)

Skin reactions assessed at 24 hours after application

10% w/v

1% w/v

0.1% w/v

1

2

3

4

5

6

7 control

8 control

9 control

10 control

410

410

430

321

308

318

340

355

344

351

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

 

Interpretation of results:
GHS criteria not met
Conclusions:
No erythema was noted in any of the test or control animals. The test article was not considered to be a strong sensitiser.
Executive summary:

A study was carried out to determine the skin-sensitising potential of test article in the albino guinea pig. The study was performed in accordance with Standard Operating Procedures.

 

Ten young female Hartley-Dunkin albino guinea pigs were used for this study. They were conditioned to the laboratory environment for not less than 6 days and on the day before initial treatment they were weighed. At this time the body weights of the guinea pigs were within the range 300 - 430 g. Three animals were marginally over the body weight range required by the protocol.

 

The test article (0.1 ml of a 10% w/v solution in dimethylformamjde) was applied daily by means of a glass syringe to the outer surface of the ears of 6 guinea pigs (animal numbers 1 - 6) for three days (days 1, 2 and 3).

On Day 8, 0.2 ml of the challenge solutions (10% w/v, 1% w/v and 0.1% w/v in dimethylformamide) was applied topically to 1 cm diameter circular areas on the clipped flanks of each of the same 6 animals. Solutions of test article were also applied in the same way on Day 8 to the clipped flanks of control animals (animal numbers 7 - 10) which had no previous treatment on the ears. The applications were made on both flanks of all 10 guinea pigs, with each concentration being applied to each flank.

The highest concentration was applied closest to the posterior end of the animal while the lowest concentration was applied nearest the anterior end.

The erythema produced on each site was assessed 24 hours later (Day 9).

 

Results

No erythema was noted in any of the test or control animals. The test article was not considered to be a strong sensitiser.

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

A study was carried out to determine the skin-sensitising potential of test article in the albino guinea pig. The study was performed in accordance with Standard Operating Procedures.

Ten young female Hartley-Dunkin albino guinea pigs were used for this study. They were conditioned to the laboratory environment for not less than 6 days and on the day before initial treatment they were weighed. At this time the body weights of the guinea pigs were within the range 300 - 430 g. Three animals were marginally over the body weight range required by the protocol.

The test article (0.1 ml of a 10% w/v solution in dimethylformamjde) was applied daily by means of a glass syringe to the outer surface of the ears of 6 guinea pigs (animal numbers 1 - 6) for three days (days 1, 2 and 3).

On Day 8, 0.2 ml of the challenge solutions (10% w/v, 1% w/v and 0.1% w/v in dimethylformamide) was applied topically to 1 cm diameter circular areas on the clipped flanks of each of the same 6 animals. Solutions of test article were also applied in the same way on Day 8 to the clipped flanks of control animals (animal numbers 7 - 10) which had no previous treatment on the ears. The applications were made on both flanks of all 10 guinea pigs, with each concentration being applied to each flank.

The highest concentration was applied closest to the posterior end of the animal while the lowest concentration was applied nearest the anterior end. The erythema produced on each site was assessed 24 hours later (Day 9).

No erythema was noted in any of the test or control animals. The test article was considered not to be a strong sensitiser.

A study was conducted to determine the skin sensitisation potential of the structural analogue according to OECD Guideline 429 (local lymph node assay) in compliance with GLP. Male CBA/Ca mice were exposed to the test substance at concentrations of 0, 3, 10 and 30% in acetone. Isotope (3H-methyl thymidine) incorporation was measured in lymph nodes. The negative (vehicle alone) and positive (hexylcinnamaldehyde at concentrations of 1, 3, and 10% w/v in acetone) controls were valid. The isotope incorporation was increased by less than a threefold at all concentrations of the test substance. Under the study conditions, the test substance was considered to be not sensitizing to mouse skin.

Respiratory sensitisation

Endpoint conclusion
Endpoint conclusion:
no study available

Justification for classification or non-classification

The results of the above studies do not trigger a classification for skin sensitisation according to CLP (EC 1272/2008) criteria.