Registration Dossier

Administrative data

Description of key information

- OECD 429; GLP; CBA mice; 1%, 3%, 10%; skin sensitizer (2004)

- OECD 429; GLP; CBA mice; 3%, 10%, 30%; skin sensitizer (2001)

- OECD 406; GLP; Guinea-pigs; 0.25%, 15%, 30%; skin sensitizer (1995)

- Equivalent to OECD 406; GLP not specified; Guinea-pigs; 1%, 2.5%, 5%, 8.5%; skin sensitizer (1983)

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:
weight of evidence
Study period:
Nov 2003 - Jan 2004
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)
GLP compliance:
yes (incl. certificate)
Type of study:
mouse local lymph node assay (LLNA)
Species:
mouse
Strain:
CBA
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Harlan Winkelmann GmbH, Borchen, Germany
- Age at study initiation: about 6-8 weeks
- Weight at study initiation: 17.3 - 20.0 g
- Housing: individually in Makrolon type I cages
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: 15 days before the first test substance application


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-24 °C
- Humidity (%): 30-70 %
- Photoperiod (hrs dark / hrs light): 12/12
Vehicle:
other: acetone
Concentration:
0%, 1%, 3% and 10% in acetone
No. of animals per dose:
6
Details on study design:
MAIN STUDY
ANIMAL ASSIGNMENT AND TREATMENT
The animals were assigned randomly to the groups and cages using randomization instructions
- Name of test method: Local Lymph Node Assay
- Criteria used to consider a positive response:
In order to reveal a possible induction of sensitization, the response in the draining lymph
node after epicutaneous application of several concentrations of the test substance to the
skin of the ear backs is determined. The parameters used to characterize the response are
lymph node weight and cell count measured. Because not only sensitization induction
but also irritation of the ear skin by the test substance may induce a lymph node responses, the
weight of ear punches taken from the area of test substance application is determiried as a
parameter for inflammatory ear swelling serving as an indicator for the irritant action of the
test substance.
If a test substance does not show a statistically significant and/or biologically relevant
increase in cell count and/or lymph node weight as compared to the vehicle control in the
presence of statistically significantly and/or biologically relevant increased ear weights as
indication of skin irritation, it is considered not to be a sensitizer .
If at least one concentration tested causes a concentration dependent statistically significant
and/or biologically relevant increase in cell count and/or lymph node weight without being
accompanied by a statistically significant and/or biologically relevant increase in ear weight,
the test substance is considered to be a sensitizer.
If statistically significant and/or biologically relevant increases in ear weights are running in
parallel to the increase in cell count and/or lymph node weight, it cannot be ruled out, that the
lymph node response was caused by irritation and not by skin sensitization. Then, for
identification of the relevance of the statistical evaluation, a comparison of the results of the
present test to appropriate historical control values is performed. If one or a
combination of the measured parameters change statistical significance, evaluation on basis
of the criteria described above may be possible. If the statistical comparison with the
historical control does not yield results useful for evaluation, further investigations may be
necessary to differentiate between irritation and sensitization response.
If a test substance does not elicit a statistical significant increase in lymph node weight and/or
cell count but shows a clear concentration related increase in response, further investigating
of the sensitization potential at higher concentrations should be considered.

TREATMENT PREPARATION AND ADMINISTRATION:
Test substance preparations were produced on a weight by weight (w/w) basis shortly before the appliacation by stirring with a magnetic stirrer.
Vehicle: Acetone
Reason for the vehicle : Acetone was used as the vehicle because good solubility of the preparation was achieved.
Form of application: Solution, epicutaneous application is simulating dermal contact with the compound which is possible to occur under practical use
conditions.
Application volume: 25 μl per ear
Site of application : Dorsal part of both ears
Positive control substance(s):
other: not included in the study
Statistics:
Mean values and standard deviations of the measured parameters were calculated for the test and control groups from the individual values.
The indices of lymph node weight, cell count and ear weight were calculated as the ratio of the test group mean values for these parameters divided by those of the vehicle control group. Lymph node weight, cell count and ear weight were statistically analyzed by Wilcoxon-test.
Positive control results:
A concurrent positive control (reliability check) with a known sensitizer was not included into this study.
Studies using the positive control substance Alpha-Hexylcinnamaldehyde techn. 85% are performed twice a year in the laboratory in order to show that the test system is able to detect sensitizing compounds under the test conditions chosen.
Parameter:
other: Cell count index
Value:
>= 2.05 - <= 3.8
Test group / Remarks:
1%, 3%, 10%
Parameter:
other: Lymph node weight index
Value:
>= 1.56 - <= 2.57
Test group / Remarks:
1%, 3%, 10%
Parameter:
other: Ear weight index
Value:
>= 1.06 - <= 1.89
Test group / Remarks:
1%, 3%, 10%
Parameter:
SI
Remarks on result:
not measured/tested
Cellular proliferation data / Observations:
Treatment of the mice with 1%, 3% and 10% test substance preparations induced statistically significant increase in lymph node cell counts as compared to the vehicle control group. The lymph node weights were statistically increased and therefore in congruency with the cell counts. A concentration < 1% in acetone is considered to be the threshold of sensitization induction and a 1 % concentration in acetone is considered to be the threshold of irritation in the present test model.

Treatment

Lymph node weight index

Cell count index

Ear weight index

Vehicle acetone

1.00

1.00

1.00

1 % in acetone

1.56**

2.05**

1.06

3 % in acetone

2.40**

3.02**

1.21**

10 % in acetone

2.57**

3.80**

1.89**

The statistical evaluations were performed using the Wilcoxon-test (* p<0.05; ** p<0.01)

 

 

 

 

No signs of systemic toxicity were noticed. The test substance induced a statistically significant and biologically relevant response of the auricular lymph nodes when applied as 1%, 3% or 10% preparations in acetone. The statistically significant increase in ear weight indicates irritation of the ear skin at concentrations of 3 and 10%, which might have interfered with the lymph node response. The strong increase in ear weight, which was accompanied by crust formation, however, indicated a significant irritant property of the test substance, when applied in these concentrations. The 1% test substance preparation in acetone induced a statistically significant increase in cell count, but only a marginal increase in ear weight. Therefore a concentration < 1% in acetone is considered to be the threshold of sensitization induction and a 1% concentration in acetone is considered to be the threshold of irritation in the present test model.

Thus it is concluded that Tripropylenglykoldiacrylat has a skin sensitizing effect in the Murine Local Lymph Node Assay under the test conditions chosen.

 

 

Interpretation of results:
Category 1 (skin sensitising) based on GHS criteria
Conclusions:
The test substance has a skin sensitizing effect in the Murine Local Lymph Node Assay under the test conditions chosen.
Executive summary:

The skin sensitizing potential of the test substance was assessed using the Murine Local Lymph Node Assay. The assay simulates the induction phase for skin sensitization in mice. It determines the response of the auricular lymph nodes on repeated application of the test substance to the dorsal skin of the ears.

Groups of 6 female CBA/Ca mice each were treated with 1%, 3% and 10% w/w preparations of the test substance in Acetone or with the vehicle alone. The study used 3 test groups and 1 control group . Each test animal was applied with 25 µ1 per ear of the respective test substance preparation to the dorsum of both ears for three consecutive days. The control group was treated with 25 µ1 per ear of the vehicle alone. Three days after the last application the mice were sacrificed and the auricular lymph nodes were removed. Lymph node response was evaluated by measuring the cellular content (indicator of cell proliferation) and weight of each animal's pooled lymph nodes. Moreover, a defined area with a diameter of 0.8 cm was punched out of the apical part of each ear and for each animal the weight of the pooled punches was determined in order to obtain an indication of possible skin irritation.

The mean indices (fold of change as compared to the vehicle control) for lymph node weight and cell count and ear weight were 1.56, 2.40, 2.57, and 2.05, 3.02, 3.80, and 1.06, 1.21, 1.89 for 1%, 3%, and 10%, resspectively.

No signs of systemic toxicity were noticed. The test substance induced a statistically significant and biologically relevant response of the auricular lymph nodes when applied as 1%, 3% or 10% preparations in acetone. The statistically significant increase in ear weight indicates irritation of the ear skin at concentrations of 3 and 10%, which might have interfered with the lymph node response . The strong increase in ear weight, which was accompanied by crust formation, however, indicated a significant irritant property of the test substance, when applied in these concentrations . The 1% test substance preparation in acetone induced a statistically significant increase in cell count, but only a marginal increase in ear weight . Therefore a concentration < 1% in acetone is considered to be the treshold of sensitization induction and a 1% concentration in acetone is considered to be the treshold of irritation in the present test model.

Thus, it is concluded that the test substance has a skin sensitizing effect in the Murine Local Lymph Node Assay under the test conditions chosen.

Endpoint:
skin sensitisation: in vivo (LLNA)
Type of information:
experimental study
Adequacy of study:
weight of evidence
Study period:
Mar 2000 - May 2000
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 406 (Skin Sensitisation)
GLP compliance:
yes
Type of study:
mouse local lymph node assay (LLNA)
Specific details on test material used for the study:
- Physical state: liquid
- Analytical purity: 82.9%
- Lot/batch No.: CEFIC 07032000
- Storage condition of test material: Refrigerator in the dark
Species:
mouse
Strain:
CBA
Sex:
male
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Harlan UK
- Age at study initiation: young adults
- Housing: a maximum of 4 mice was housed per cage
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: at least 5 days


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22+-3 °C
- Humidity (%): 30-70%
- Air changes (per hr): a minimum of 15 changes per hour
- Photoperiod (hrs dark / hrs light): 12/12
Vehicle:
other: acetone
Concentration:
0%, 3%, 10% and 30% w/v preparations of the test substance in acetone
No. of animals per dose:
4
Details on study design:
MAIN STUDY
ANIMAL ASSIGNMENT AND TREATMENT
- Name of test method: Local Lymp Node Assay
- Criteria used to consider a positive response: 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 moderate or greater responses in standard guinea pig tests for skin sensitisation.
Consequently, a test substance which does not fulfill the above criterion is designated as unlikely to be a moderate or strong sensitiser.
TREATMENT PREPARATION AND ADMINISTRATION:
Groups of four male mice were used for this study. Approximately 25μl 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.
Positive control substance(s):
hexyl cinnamic aldehyde (CAS No 101-86-0)
Statistics:
no data
Positive control results:
In a positive control study, hexylcinnamaldehyde was shown to have the capacity to cause skin sensitization when applied as 3% or 10% w/v preparations in acetone, confirming the validity of the protocol used for this study.
Parameter:
SI
Value:
14.83
Test group / Remarks:
3%
Remarks on result:
other: (3 %)
Parameter:
SI
Value:
12.88
Test group / Remarks:
10%
Remarks on result:
other: (10 %)
Parameter:
SI
Value:
17.91
Test group / Remarks:
30%
Remarks on result:
other: (30 %)
Parameter:
other: disintegrations per minute (DPM)
Value:
>= 57 616 - <= 80 096
Test group / Remarks:
3%, 10%, 30%
Remarks on result:
other: control: 4473 dpm; 3% solution: 66331 dpm, 10% solution 57616 dpm, 30% solution: 80096 dpm

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 greater than 3-fold at all three concentrations. Consequently, the test substance was shown to be a potential skin sensitiser. Following, the third application, the ears of animals dosed with the 30% w/v preparation were red and swollen.

Interpretation of results:
Category 1 (skin sensitising) based on GHS criteria
Conclusions:
In conclusion, the test substance is a skin sensitiser under the conditions of the test. However, taking into account the observed irritating effects on the ears, and the missing dose-dependency of the lymph node proliferation, the sensitising effect cannot be clearly discriminated from at least partly possible lymph node proliferation based on skin irritancy.
Executive summary:

A sample of the test substance was assessed for its skin sensitisation potential using tlie mouse Local Lymph Node Assay . The assay determines the level of T lymphocyte proliferation in the lymph nodes draining the site of chemical application, by measuring the amount of radiolabelled thymidine incorporated into the dividing cells . The test substance was applied as 3%, 10% or 30% w/v preparations in acetone.

The test substance was shown to have the capacity to cause skin sensitisation when applied as 3%, 10% or 30% w/v preparations in acetone. In a positive control study, hexylcinnamaldehyde was shown to have the capacity to cause skin sensitization when applied as 3% or 10% w/v preparations in acetone, confirmirig the validity of the protocol used for this study.

In conclusion, the test substance is a skin sensitiser under the conditions of the test. However, taking into account the observed irritating effects on the ears, and the missing dose-dependency of the lymph node proliferation, the sensitising effect cannot be clearly discriminated from at least partly possible lymph node proliferation based on skin irritancy .

Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
experimental study
Adequacy of study:
weight of evidence
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Qualifier:
according to
Guideline:
EU Method B.6 (Skin Sensitisation)
GLP compliance:
yes
Type of study:
guinea pig maximisation test
Justification for non-LLNA method:
At the time the study was conducted, there was not existing guideline for the LLNA method.
Specific details on test material used for the study:
- Physical state: clear colourless liquid
- Analytical purity: > 98 %
- Lot/batch No.: YJA 704
- Storage condition of test material: at room temperature in the dark
Species:
guinea pig
Strain:
Dunkin-Hartley
Sex:
male
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: D. Hall, Newchurch, UK
- Age at study initiation: approx. 4-5 weeks
- Weight at study initiation: 284-343 g
- Housing: in groups of 5 in suspended metal cages with wire mesh floors
- Diet: Vitamin C enriched guinea pig diet FD2, ad libitum
- Water: ad libitum
- Acclimation period: 5 days

ENVIRONMENTAL CONDITIONS
- Temperature: approx. 21 °C
- Humidity: 30 - 70 %
- Air changes: approx. 15 per hr
- Photoperiod: 12 hrs dark / 12 hrs light
Route:
intradermal and epicutaneous
Vehicle:
other: Alembicol D
Concentration / amount:
Preliminary study:
Induction:
intradermal injection: 0.25 % v/v in Alembicol D
topical application: as supplied (no vehicle)
Challenge:
topical: 30 and 15 % v/v in Alembicol D

Main Study:
Induction:
Intradermal induction:
Control animals: (1) 0.1 ml of Freund's complete adjuvant 50 : 50 with sterile water for irrigation (Ph. Eur.). (2) 0.1 ml of Alembicol D. (3) 0.1 ml of Freund's complete adjuvant 50 : 50 with Alembicol D.
Test animals: (1) 0.1 ml of Freund's complete adjuvant 50 : 50 with sterile water for irrigation (Ph. Eur.). (2) 0.1 ml of Sartomer 306 Monomer/TPGDA, 0.25 % v/v in Alembicol D. (3) 0.1 ml of Sartomer 306 Monomer/TPGDA, 0.25 % v/v in a 50: 50 mixture of Alembicol D and Freund's complete adjuvant.
A volume of 0.1 ml was injected into both the left and right injection sites.
Challenge:
topical application: 30 and 15 % v/v in Alembicol D (two weeks after the topical induction)
Route:
epicutaneous, occlusive
Vehicle:
other: Alembicol D
Concentration / amount:
Preliminary study:
Induction:
intradermal injection: 0.25 % v/v in Alembicol D
topical application: as supplied (no vehicle)
Challenge:
topical: 30 and 15 % v/v in Alembicol D

Main Study:
Induction:
Intradermal induction:
Control animals: (1) 0.1 ml of Freund's complete adjuvant 50 : 50 with sterile water for irrigation (Ph. Eur.). (2) 0.1 ml of Alembicol D. (3) 0.1 ml of Freund's complete adjuvant 50 : 50 with Alembicol D.
Test animals: (1) 0.1 ml of Freund's complete adjuvant 50 : 50 with sterile water for irrigation (Ph. Eur.). (2) 0.1 ml of Sartomer 306 Monomer/TPGDA, 0.25 % v/v in Alembicol D. (3) 0.1 ml of Sartomer 306 Monomer/TPGDA, 0.25 % v/v in a 50: 50 mixture of Alembicol D and Freund's complete adjuvant.
A volume of 0.1 ml was injected into both the left and right injection sites.
Challenge:
topical application: 30 and 15 % v/v in Alembicol D (two weeks after the topical induction)
No. of animals per dose:
15 animals: 5 controls and 10 treated with the test material of both concentrations
Details on study design:
Induction:
- Intradermal induction:
A 40 mm x 60 mm area of dorsal skin on the scapular region of the guinea-pig was clipped free of hair with electric clippers. Three pairs of intradermal injections were made into a 20 mm x 40 mm area within the clipped area.
-Topical application:
One week after the injections, the same 40 x 60 mm interscapular area was clipped and shaved free of hair.
A 20 x 40 mm patch of Whatman No. 3 paper was saturated with approximately 0.4 ml of Sartomer 306 Monomer/TPGDA, as supplied. The patch was placed on the skin of the test animals and covered by a length of impermeable plastic adhesive tape (50 mm width "Blenderm"). This in turn was firmly secured by elastic adhesive bandage (50 mm width "Elastoplast") wound round the torso of the animal and fixed with "Sleek" impervious plastic
adhesive tape. The dressing was left in place for 48 hours.
During the induction phase, the control animals were treated similarly to the test animals with the exception that the test substance was omitted from the intradermal injections and topical application.

Challenge - control and test animals:
The control and test animals were challenged topically two weeks after the topical induction application using Sartomer 306 Monomer/TPGDA, 30 and 15% v/v in Alembicol D. Hair was removed by clipping and then shaving from an area on the left flank of each guinea-pig. A 20 mm x 20 mm patch of Whatman No. 3 paper was saturated with approximately 0.2 ml of Sartomer 306 Monomer/TPGDA, 30% v/v in Alembicol D and applied to an anterior site on the flank. Sartomer 306 Monomer/TPGDA, 15% v/v in Alembicol D was applied in a similar manner to a posterior site. The patches were sealed to the flank for 24 hours under strips of "Blenderm" covered by 11 Elastoplast" wound raund the trunk and secured with "Sleek".

OBSERVATIONS
Clinical signs:
All animals were observed daily for signs of ill health or toxicity.

Bodyweight:
The bodyweight of each guinea-pig on the main study was recorded on Day 1 (day of intradermal injections) and on the last day observations were made of dermal responses to the challenge applications.

Dermal responses:
The dermal reactions resulting from intradermal injection and topical application on the preliminary study, and topical application at the challenge were assessed using the numerical scoring system by Draize.
Positive control substance(s):
not required
Remarks:
The sensitivity of the guinea-pig strain used is checked periodically with hexyl cinnamic aldehyde, a known sensitiser.
Positive control results:
The sensitivity of the guinea-pig strain used is checked periodically with hexyl cinnamic aldehyde, a known sensitiser.
Reading:
1st reading
Hours after challenge:
24
Group:
negative control
Dose level:
15 %
No. with + reactions:
0
Total no. in group:
5
Reading:
1st reading
Hours after challenge:
24
Group:
negative control
Dose level:
30 %
No. with + reactions:
0
Total no. in group:
5
Reading:
1st reading
Hours after challenge:
24
Group:
test group
Dose level:
15 %
No. with + reactions:
2
Total no. in group:
10
Clinical observations:
Localized dermal reaction in one animal
Remarks on result:
other: Number with positive reactions are not tantamount to a positive indication of sensitization but also to irritation.
Reading:
1st reading
Hours after challenge:
24
Group:
test group
Dose level:
30 %
No. with + reactions:
8
Total no. in group:
10
Clinical observations:
Blanching; necrotic patch; localized dermal reaction in one animal each
Remarks on result:
other: Number with positive reactions are not tantamount to a positive indication of sensitization but also to irritation.
Reading:
2nd reading
Hours after challenge:
48
Group:
negative control
Dose level:
15 %
No. with + reactions:
0
Total no. in group:
5
Reading:
2nd reading
Hours after challenge:
48
Group:
negative control
Dose level:
30 %
No. with + reactions:
1
Total no. in group:
5
Reading:
2nd reading
Hours after challenge:
48
Group:
test group
Dose level:
15 %
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
Dryness and sloughing of the epidermis in one animal
Reading:
2nd reading
Hours after challenge:
48
Group:
test group
Dose level:
30 %
No. with + reactions:
7
Total no. in group:
10
Clinical observations:
Thickening, dryness and sloughing of the epidermis; necrotic patch; localized dermal reaction in one animal each
Remarks on result:
other: Number with positive reactions are not tantamount to a positive indication of sensitization but also to irritation.
Reading:
other: 3rd reading
Hours after challenge:
72
Group:
negative control
Dose level:
15 %
No. with + reactions:
0
Total no. in group:
5
Reading:
other: 3rd reading
Hours after challenge:
72
Group:
negative control
Dose level:
30 %
No. with + reactions:
0
Total no. in group:
5
Clinical observations:
Dryness and sloughing of the epidermis; necrotic patch in one animal each
Reading:
other: 3rd reading
Hours after challenge:
72
Group:
test group
Dose level:
15 %
No. with + reactions:
0
Total no. in group:
10
Clinical observations:
Dryness and sloughing of the epidermis in 3 animals
Reading:
other: 3rd reading
Hours after challenge:
72
Group:
test group
Dose level:
30 %
No. with + reactions:
8
Total no. in group:
10
Clinical observations:
Thickening, dryness and sloughing of the epidermis in 4 animals; localized dermal reaction in 3 animals; dryness and sloughing of the epidermis in 2 animals; necrotic patch in 1 animal
Remarks on result:
other: Number with positive reactions are not tantamount to a positive indication of sensitization but also to irritation.
Reading:
1st reading
Group:
positive control
Dose level:
As supplied
No. with + reactions:
10
Total no. in group:
10
Remarks on result:
other: checked periodically and not included in this study

Table 1: Dermal reactions observed after the challenge application with the test substance

Freud's treated control

 

 

Guinea-pig number

 

E=Erythema

0 =Oedema

Score

24

Hours

48

Hours

72 Hours

A

p

A

p

A

p

1800

E

0

0

0

0

0

0

0

0

0

 

0

O*

0

0

1801

E

0

0

0

0

0

10

0

0

 

NP1

0

0

0

 

1802

E

0

0

0

0

0

0

0

0

0

 

0

0

0

0

1803

E

0

0

0

0

0

0

0

0

0

 

0

0

0

0

1804

E

0

0

0

0

0

0

0

0

0

 

0

0

0

0

 

NP Necrotic patch

* Dryness and sloughing of the epidermis

AAnterior site, exposed to the test substance, 30% v/v in Alembicol D

P Posterior site. exposed to the test substance, 15 % v/v in Alembicol D

 

 Test animals

 

 

 

Score

 

 

 

24 Hours

48 Hours

72 Hours

 

Guinea-pig number

E = Erythema

0 = Oedema

A

P

A

P

A

P

Results Positive ( +) Negative(-)

Inconclusive ( ±)

1805

E

O

1

0

0

0

1

0

0

0

1

0

0

0*

-

1806

E

O

0

0

0

0

0

0

0

0

0

0

0

0

-

1807

E

O

0

0

L1

0

0

0

0

0

ØL1

0

0

0

±

1808

E

O

1

0

0

0

0

0

0

0

0

0*

0

0*

-

1809

E

O

#1

1

0

0

1

1

0

0

Ø1

1

0

0

+

1810

E

O

1

1

0

0

Ø1

1

0

0*

Ø1

1

0

0*

+

1811

E

O

1

0

0

0

1

1

0

0

Ø1

1

0

0

+

1812

E

O

NP1

0

0

0

NP1

1

0

0

ØNP1

1

0

0

+

1813

E

O

L1

0

0

0

L1

0

0

0

L1

0*

0

0

-

1814

E

O

1

1

1

0

1

0

0

0

L1

0

0

0

-

 

 

 

LLocalised dermal reaction (restricted to a small area of the challenge site)

NP Necrotic patch

* Dryness and sloughing of the epidermis

ØThickening, dryness and sloughing of the epidermis

A Anterior site, exposed to the test substance, 30% v/v in Alembicol D

P Posterior site. exposed to the test substance, 15% v/v in Alembicol D

# Blanching

CLINICAL SIGNS

No signs of ill health or toxicity were recorded.

BODYWEIGHT

Bodyweight increases were recorded for all guinea-pigs over the period of the study.

INDUCTION

Intradermal injections: Necrosis was recorded at sites receiving Freund's Complete Adjuvant in test and control animals. Slight irritation was seen in test animals at sites receiving the test substance, 0.25 % v/v in Alembicol D and slight irritation was observed in control animals receiving Alembicol D alone.

Topical application: Moderate erythema was observed in test animals following topical application with the test substance, as supplied and no erythema was seen in the control guinea-pigs.

Interpretation of results:
Category 1 (skin sensitising) based on GHS criteria
Conclusions:
Under the experimental conditions chosen, the test material caused sensitization reactions after challenge on the skin of guinea pigs. Therefore, the test substance fulfills the requirements to be classified as skin sensitiser category 1 according to GHS criteria.
Executive summary:

A study was performed to assess the skin sensitisation potential of the test substance using the guinea-pig. Based on the results of a preliminary study and in compliance with the guideline, the following close levels were selected:

- Intradermal injection: 0.25% v/v in Alembicol D

- Topical application: as supplied

- Challenge application: 30 and 15% v/v in Alembicol D

Ten test and five control guinea-pigs were used in this study.

In this study, the test substance produced evidence of skin sensitisation (delayed contact hypersensitivity) in four of the ten test animals. A further one animal gave an inconclusive response. The remaining five animals gave a negative response.

Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
experimental study
Adequacy of study:
weight of evidence
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
study well documented, meets generally accepted scientific principles, acceptable for assessment
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 406 (Skin Sensitisation)
GLP compliance:
not specified
Type of study:
guinea pig maximisation test
Justification for non-LLNA method:
The test was performed prior to the implementation of the LLNA test method.
Species:
guinea pig
Strain:
Hartley
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Connaught Laboratories, Toronto, Canada
- Weight at study initiation: 300-500 g
- Housing: they were housed in metal cages
- Diet: ad libitum
- Water: ad libitum



ENVIRONMENTAL CONDITIONS
no data

Route:
intradermal and epicutaneous
Vehicle:
propylene glycol
Concentration / amount:
1%, 2.5%, 5% and 8.5%
Route:
epicutaneous, occlusive
Vehicle:
propylene glycol
Concentration / amount:
1%, 2.5%, 5% and 8.5%
No. of animals per dose:
10 animals at 1%, 2.5% and 5% each, 6 animals at 8.5%
Details on study design:
see: any other information on materials and methods
Positive control substance(s):
no
Positive control results:
No positive control included.
Reading:
1st reading
Hours after challenge:
48
Group:
test group
Dose level:
1.0 %
No. with + reactions:
3
Total no. in group:
10
Reading:
1st reading
Hours after challenge:
48
Group:
test group
Dose level:
2.5 %
No. with + reactions:
4
Total no. in group:
10
Reading:
1st reading
Hours after challenge:
48
Group:
test group
Dose level:
5 %
No. with + reactions:
5
Total no. in group:
10
Reading:
1st reading
Hours after challenge:
48
Group:
test group
Dose level:
8.5 %
No. with + reactions:
3
Total no. in group:
6
Group:
negative control
Remarks on result:
not measured/tested
Group:
positive control
Remarks on result:
not measured/tested

Probit analysis was carried out using log-probability paper to calculate the intradermal (ID) concentration required to sensitise half the guineapigs (IDSC50) The IDSC50 for the test substance was 3.5%.

Interpretation of results:
Category 1B (indication of skin sensitising potential) based on GHS criteria
Conclusions:
Under the experimental conditions chosen, the test material caused sensitization reactions after challenge on the skin of guinea pigs. Therefore, classification as skin sensitizer Cat. 1B is required.
Executive summary:

The study was performed equivalent to OECD 406 and with no information concerning GLP compliance (1983b). Hartley guinea pigs were induced in two stages; in the first stage either side of the shoulder region was injected intradermally. One week after injection, the shoulder region of each animal was clipped. The chemical was applied in its respective concentrations (1, 2.5, 5 or 8.5%) in petrolatum over the sites of previous intradermal injection for 48 hours. All animals were challenged two weeks after topical exposure using nonirritant concentrations that had been determined in control animals. Patches were applied to assess irritancy. After 24 hours, patches were removed. Sites were examined at 48 hours for evidence of reaction. Sensitizing effects were seen at concentrations of 1, 2.5, 5 and 8.5% in 3/10, 4/10, 5/10 and 3/6 animals, respectively. Thus, the test substance was considered to be a skin sensitizer.

Endpoint conclusion
Endpoint conclusion:
adverse effect observed (sensitising)
Additional information:

There are valid in vivo data available for the assessment of the skin sensitization potential of the test substance presented as weight of evidence.

 

In a Local Lymph Node Assay, acc. to OECD 429 and in compliance with GLP, six female CBA mice per dose were treated with 0, 1, 3 and 10 %, of the test material diluted in acetone under standard conditions (2004). Twenty-five microliter were applied to the dorsum of each ear for three consecutive days. The control group was treated with 25 µl per ear of the vehicle alone. Three days after the last application the mice were sacrificed, and the auricular lymph nodes were removed. The lymph node response was evaluated by measuring the cellular content (indicator of cell proliferation) and weight of each animal's pooled lymph nodes. Moreover, a defined area with a diameter of 0.8 cm was punched out of the apical part of each ear and for each animal the weight of the pooled punches was determined in order to obtain an indication of possible skin irritation. No signs of systemic toxicity were noticed. A statistically significant increase in ear weight as an indication of skin irritation was observed at concentrations of 3 and 10%. A simultaneous response to irritation is indistinguishable from a sensitization response in this test system. The 1% test substance preparation in acetone induced a statistically significant increase in cell count, but only a marginal increase in ear weight. Therefore, a concentration < 1% in acetone is considered to be the threshold of sensitization induction and a 1% concentration in acetone is considered to be the threshold of irritation in the present test model.

 

The sensitizing properties of the test material were also shown in a second LLNA, acc. to OECD 429 and in compliance with GLP (2001).Groups of four CBA 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 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. 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 greater than 3-fold at all three concentrations. Following, the third application, the ears of animals dosed with the 30% w/v preparation were red and swollen.

 

GPMTs were conducted in two further studies. The first study was performed equivalent to OECD 406 and with no information concerning GLP compliance (1983b). Hartley guinea pigs were induced in two stages; in the first stage either side of the shoulder region was injected intradermally. One week after injection, the shoulder region of each animal was clipped. The chemical was applied in its respective concentrations (1, 2.5, 5 or 8.5%) in petrolatum over the sites of previous intradermal injection for 48 hours. All animals were challenged two weeks after topical exposure using nonirritant concentrations that had been determined in control animals. Patches were applied to assess irritancy. After 24 hours, patches were removed. Sites were examined at 48 hours for evidence of reaction. Sensitizing effects were seen at concentrations of 1, 2.5, 5 and 8.5% in 3/10, 4/10, 5/10 and 3/6 animals, respectively. Thus, the test substance was considered to be a skin sensitizer.

In the second GPMT, fifteen Dunkin-Hartley guinea pigs (5 controls, 10 substance) were treated with the test material. The study was performed according to OECD 406 and in compliance with GLP regulations (1995). Based on the results of a preliminary study and in compliance with the guideline, three dose levels were selected. For intradermal injection, 0.25% v/v in Alembicol D were utilized and for the topical application, the test substance was used as supplied. Challenge applications were performed with the test substance of 30 and 15% v/v in Alembicol D two weeks after the topical induction. Ten test and five control guinea-pigs were used in this study. In this study, the test substance produced evidence of skin sensitisation (delayed contact hypersensitivity) in four of the ten test animals. A further one animal gave an inconclusive response. The remaining five animals gave a negative response.

 

In another study, which was not compliant with GLP regulations, contact skin reactions were investigated using guinea-pigs that were immunized by the Polak method (1983a). On day 7, the animals received 4 footpad injections of 0.1 ml of an emulsion containing 2 mg/ml of the chemical, in ethanol:saline (1:4), in Freund's complete adjuvant (FCA - Difco mycobacterium butyricum). In addition, 0.1 ml of the emulsion was injected into the nape of the neck . The guinea pigs received a total of 1 mg of chemical. On day 7, open skin testing was performed by dropping 0.02 ml of a solution of the chemical in acetone:olive oil (4:1) onto the shaved flank. The concentration of the skin test solution varied with the chemical and those used were dilutions of 5% or the maximum concentration which gave no non-specific irritation. Skin tests were repeated weekly at different sites on the flank for up to 12 weeks. The skin test concentrations were 0.4% and 1.1%. The first positive skin test was observed after 28 days. The test substance was judged to be a weak sensitizer.

 

For the sensitising potential of the test substance, a qualitative assessment was conducted:

 

Although data from local lymphnode assays is available the derivation of an EC3 was not possible. The available LLNA data indicates, that the EC3 is definitely below 1%.

The available data from the guinea pig tests indicates that the intradermal induction concentration was always above or equal to 0.25 % leading to animals’ responses between 30% and 80% varying from study to study.

Respiratory sensitisation

Endpoint conclusion
Endpoint conclusion:
no study available

Justification for classification or non-classification

The available experimental test data are reliable and suitable for classification purposes under Regulation (EC) 1272/2008. The test substance was shown to be a skin sensitizer. In addition, it has also been legally classified as skin sensitizer (Annex VI, 1272/2008), and is therefore classified as Skin sensitizing Category 1 according to GHS-criteria.

 

There are no data available to classify tripropylene glycol diacrylate as a sensitizer of the respiratory tract.