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Diss Factsheets

Administrative data

Description of key information

The substance was not skin sensitising in a LLNA.

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:
key study
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)
GLP compliance:
yes (incl. QA statement)
Type of study:
mouse local lymph node assay (LLNA)
Specific details on test material used for the study:
SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: A01297 (Cinilex DPP RED SR1C)
- Expiration date of the lot/batch: -
- Purity test date: -

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: room tempreature in the dark

Species:
mouse
Strain:
CBA
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Harlan UK Limited, Bicester, Oxon, UK
- Housing: individually
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19 -25 °C
- Humidity (%): 30-70%
- Air changes (per hr): 15/h
- Photoperiod (hrs dark / hrs light): 12/12

Vehicle:
acetone/olive oil (4:1 v/v)
Concentration:
5%, 10% or 25% w/w in acetone/olive oil 4:1
No. of animals per dose:
4
Key result
Parameter:
SI
Value:
0.99
Test group / Remarks:
5%
Parameter:
SI
Test group / Remarks:
10%
Remarks on result:
other: no results due to accidential spillage of pooled lymph node cell suspension
Key result
Parameter:
SI
Value:
1.13
Test group / Remarks:
25%
Parameter:
other: disintegrations per minute (DPM)
Remarks on result:
other: vehicle: 7694.44 5%: 7593.42 25% 8718.31

Due to accidental spillage of the single cell suspension of pooled lymph node cells for the group treated with the test material at a concentration of 10%, the results obtained cannot be verified as being accurate. For this reason the results for the 10% group were not used for reporting purposes. This was considered not to affect the purpose or integrity of the study.

Interpretation of results:
GHS criteria not met
Conclusions:
In a GLP-study according to OECD test guideline 429 (LLNA), the substance was not skin sensitising.
Executive summary:

A study was performed to assess the skin sensitisation potential of the test material in the CBA/Ca strain mouse following topical application to the dorsal surface of the ear.

Following a preliminary screening test, three groups, each of four animals, were treated with 50 p.1 (25 p1 per ear) of the test material as a suspension in acetone/olive oil 4:1 at concentrations of 5%, 10% or 25% w/w. A further group of four animals was treated with acetone/olive oil 4:1 alone.

Due to accidental spillage of the single cell suspension of pooled lymph node cells for the group treated with the test material at a concentration of 10%, the results obtained cannot be verified as being accurate. For this reason the results for the 10% group were not used for reporting purposes. This was considered not to affect the purpose or integrity of the study.

Simulation Indexes for 5% and 25 % were 0.99 and 1.13, respectively. Therefore, the test material was considered to be a non-sensitiser under the conditions of the test.

Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Guideline study conducted according to GLP.
Qualifier:
according to guideline
Guideline:
OECD Guideline 406 (Skin Sensitisation)
Version / remarks:
(1981)
Deviations:
no
GLP compliance:
yes
Type of study:
guinea pig maximisation test
Justification for non-LLNA method:
When this study was performed, the LLNA did not yet exist as an OECD testing guideline.
Species:
guinea pig
Strain:
Dunkin-Hartley
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: CIBA-GEIGY LTD. Tierfarm, Sisseln, Switzerland
- Age at study initiation: approx. 10 weeks
- Weight at study initiation: 304 - 461 g
- Housing: individually in Macrolon type-3 cages.
- Diet (ad libitum): standard guinea pig pellets, NAFAG No. 846
- Water (ad libitum): fresh water
- Acclimation period: 9 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 +/- 3
- Humidity (%): 30 - 70
- Photoperiod (hrs dark / hrs light): 12 / 12

Route:
intradermal and epicutaneous
Vehicle:
other: For intradermal treatment sesame oil and for epidermal treatment vaseline was used as vehicle.
Concentration / amount:
Intradermal induction was performed with 1 % dilution of test article in sesame oil or in adjuvant/physiological saline. A 1:1 mixture of adjuvant and saline served as control.
Epidermal induction was conducted with 10 % test article in vaseline as a paste of 0.4 g.
The challenge was performed with 0.3 % test article in vaseline as paste of 0.2 g.


Route:
epicutaneous, occlusive
Vehicle:
other: For intradermal treatment sesame oil and for epidermal treatment vaseline was used as vehicle.
Concentration / amount:
Intradermal induction was performed with 1 % dilution of test article in sesame oil or in adjuvant/physiological saline. A 1:1 mixture of adjuvant and saline served as control.
Epidermal induction was conducted with 10 % test article in vaseline as a paste of 0.4 g.
The challenge was performed with 0.3 % test article in vaseline as paste of 0.2 g.


No. of animals per dose:
20 animals (10 males, 10 females)
Details on study design:
RANGE FINDING TESTS
The concentration of the test compound for the induction and challenge periods were determined on four separate animals with concentrations of 0.3 %, 1 %, 3 %, 10 %. No further details are given.

MAIN STUDY
A. INDUCTION EXPOSURE
- No. of exposures: 2; one intradermal injection (0.1 ml/site) of 1 % dilution in sesame oil and one epidermal application of 10 % dilution in vaseline one week after intradermal injection under occlusive conditions.
- Exposure period: Epidermal application was performed for 48 h.
- Site: neck

B. CHALLENGE EXPOSURE (TWO WEEKS AFTER INDUCTION)
- No. of exposures: 1
- Day(s) of challenge: 1
- Exposure period: 24 h
- Site: flank
- Concentrations: 0.3 % test article in vaseline
- Evaluation (hr after challenge): approximately 24 and 48 hours after challenge

SCORING
Evaluation of skin reactions according to Draize (1959)

Erythema and eschar formation
- No erythema 0
- Very slight erythema (barely perceptible) 1
- Well defined erythema 2
- Moderate to severe erythema 3
- Severe erythema (beet redness) to slight eschar formation (injuries in depth) 4

Edema formation
- No edema 0
- Very slight edema (barely perceptible) 1
- Slight edema (edges of area well defined by definite raising) 2
- Moderate edema (raised approximately 1 mm) 3
- Severe edema (raised more than 1 mm and extending beyond area of exposure) 4
Challenge controls:
During the induction period a control group was treated with adjuvant and the vehicle. One half of this group was challenged with vehicle and the other half with test article.
Positive control substance(s):
yes
Remarks:
The sensitivity of the strain is checked every six months with paraphenylene-diamine or potassium-dichromate according to Maurer and Hess 1994 (Toxicology 31, 217-222) in the testing facility.
Reading:
1st reading
Hours after challenge:
24
Group:
negative control
Dose level:
0%
No. with + reactions:
0
Total no. in group:
20
Clinical observations:
no skin irritations were observed
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 24.0. Group: negative control. Dose level: 0%. No with. + reactions: 0.0. Total no. in groups: 20.0. Clinical observations: no skin irritations were observed .
Reading:
2nd reading
Hours after challenge:
48
Group:
negative control
Dose level:
0%
No. with + reactions:
0
Total no. in group:
20
Clinical observations:
no skin irritations were observed
Remarks on result:
other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: negative control. Dose level: 0%. No with. + reactions: 0.0. Total no. in groups: 20.0. Clinical observations: no skin irritations were observed.
Reading:
1st reading
Hours after challenge:
24
Group:
test chemical
Dose level:
0.3 %
No. with + reactions:
0
Total no. in group:
20
Clinical observations:
no skin irritations were observed
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 24.0. Group: test group. Dose level: 0.3 %. No with. + reactions: 0.0. Total no. in groups: 20.0. Clinical observations: no skin irritations were observed .
Reading:
2nd reading
Hours after challenge:
48
Group:
test chemical
Dose level:
0.3 %
No. with + reactions:
0
Total no. in group:
20
Clinical observations:
no skin irritations were observed
Remarks on result:
other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: test group. Dose level: 0.3 %. No with. + reactions: 0.0. Total no. in groups: 20.0. Clinical observations: no skin irritations were observed .
Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 406 (Skin Sensitisation)
Version / remarks:
(1981)
Deviations:
no
GLP compliance:
no
Type of study:
guinea pig maximisation test
Justification for non-LLNA method:
When this study was performed, the LLNA did not yet exist as an OECD testing guideline.
Species:
guinea pig
Strain:
Dunkin-Hartley
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: CIBA-GEIGY LTD. Tierfarm, Sisseln, Switzerland
- Age at study initiation: approx. 10 weeks
- Weight at study initiation: 320 - 433 g
- Housing: individually in Macrolon type-3 cages.
- Diet (ad libitum): standard guinea pig pellets, NAFAG No. 846
- Water (ad libitum): fresh water
- Acclimation period: 8 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 +/- 3
- Humidity (%): 30 - 70
- Photoperiod (hrs dark / hrs light): 12 / 12

Route:
intradermal and epicutaneous
Vehicle:
other: For intradermal treatment sesame oil and for epidermal treatment vaseline was used as vehicle.
Concentration / amount:
Intradermal induction was performed with 1 % dilution of test article in sesame oil or in adjuvant/physiological saline. A 1:1 mixture of adjuvant and saline served as control.
Epidermal induction was conducted with 10 % test article in vaseline as a paste of 0.4 g. On day before epidermal induction the application sites were pretreated with 10 % sodium lauryl sulfate under open conditions.
The challenge was performed with 3 % test article in vaseline as paste of 0.2 g.
Route:
epicutaneous, occlusive
Vehicle:
other: For intradermal treatment sesame oil and for epidermal treatment vaseline was used as vehicle.
Concentration / amount:
Intradermal induction was performed with 1 % dilution of test article in sesame oil or in adjuvant/physiological saline. A 1:1 mixture of adjuvant and saline served as control.
Epidermal induction was conducted with 10 % test article in vaseline as a paste of 0.4 g. On day before epidermal induction the application sites were pretreated with 10 % sodium lauryl sulfate under open conditions.
The challenge was performed with 3 % test article in vaseline as paste of 0.2 g.
No. of animals per dose:
20 animals (10 males, 10 females)
Details on study design:
RANGE FINDING TESTS:
The concentration of test compound for induction and challenge periods were determined on four separate animals with concentrations of 1 %, 3 % and 10 % in vaseline.

MAIN STUDY
A. INDUCTION EXPOSURE
- No. of exposures: 2; one intradermal injection (0.1 ml/site) of 1 % dilution in sesame oil and one epidermal application of 10 % dilution in vaseline one week after intradermal injection under occlusive conditions. The application sites were pretreated with 10 % sodium lauryl sulfate (open application) one day before epidermal induction.
- Exposure period: Epidermal application was performed for 48 h.
- Site: neck

B. CHALLENGE EXPOSURE (TWO WEEKS AFTER INDUCTION)
- No. of exposures: 1
- Day(s) of challenge: 1
- Exposure period: 24 h
- Site: flank
- Concentrations: 3 % test article in vaseline
- Evaluation (hr after challenge): approximately 24 and 48 hours after challenge

SCORING
Evaluation of skin reactions according to Draize (1959)

Erythema and eschar formation
- No erythema 0
- Very slight erythema (barely perceptible) 1
- Well defined erythema 2
- Moderate to severe erythema 3
- Severe erythema (beet redness) to slight eschar formation (injuries in depth) 4

Edema formation
- No edema 0
- Very slight edema (barely perceptible) 1
- Slight edema (edges of area well defined by definite raising) 2
- Moderate edema (raised approximately 1 mm) 3
- Severe edema (raised more than 1 mm and extending beyond area of exposure) 4
Challenge controls:
During the induction period a control group was treated with adjuvant and the vehicle. One half of this group was challenged with vehicle and the other half with test article.
Positive control substance(s):
yes
Remarks:
The sensitivity of the strain is checked every six months with paraphenylene-diamine or potassium-dichromate according to Maurer and Hess 1994 (Toxicology 31, 217-222) in the testing facility.
Reading:
1st reading
Hours after challenge:
24
Group:
negative control
Dose level:
0%
No. with + reactions:
0
Total no. in group:
20
Clinical observations:
no skin irritations were observed
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 24.0. Group: negative control. Dose level: 0%. No with. + reactions: 0.0. Total no. in groups: 20.0. Clinical observations: no skin irritations were observed .
Reading:
2nd reading
Hours after challenge:
48
Group:
negative control
Dose level:
0%
No. with + reactions:
0
Total no. in group:
20
Clinical observations:
no skin irritations were observed
Remarks on result:
other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: negative control. Dose level: 0%. No with. + reactions: 0.0. Total no. in groups: 20.0. Clinical observations: no skin irritations were observed .
Reading:
1st reading
Hours after challenge:
24
Group:
test chemical
Dose level:
3 %
No. with + reactions:
0
Total no. in group:
20
Clinical observations:
no skin irritations were observed
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 24.0. Group: test group. Dose level: 3 %. No with. + reactions: 0.0. Total no. in groups: 20.0. Clinical observations: no skin irritations were observed .
Reading:
2nd reading
Hours after challenge:
48
Group:
test chemical
Dose level:
3 %
No. with + reactions:
0
Total no. in group:
20
Clinical observations:
no skin irritations were observed
Remarks on result:
other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: test group. Dose level: 3 %. No with. + reactions: 0.0. Total no. in groups: 20.0. Clinical observations: no skin irritations were observed .
Interpretation of results:
not sensitising
Remarks:
Migrated information
Endpoint:
skin sensitisation: in vivo (LLNA)
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Justification for type of information:
1. HYPOTHESIS FOR THE ANALOGUE APPROACH
The read-across is performed between two forms of the same substance. The identities of the two forms are describe below.

2. SOURCE AND TARGET CHEMICAL(S) (INCLUDING INFORMATION ON PURITY AND IMPURITIES)
The source form is 3,6-bis(4-chlorophenyl)-2,5-dihydropyrrolo[3,4-c]pyrrol-1,4-dione (EC-no. 401-540-3), referred to here as PR254. PR254 is an organic mono-constituent substance with a typical purity of > 99.5% (w/w). It does not contain any impurity relevant for classification or labelling of the substance. The target form is the nanoform of the source substance, referred to here as PR254 nanoform. As the source form, it has a typical purity of > 99.5% (w/w) and it does not contain any impurity relevant for classification or labelling of the substance. The PR254 nanoform is spheroidal with a pure polyhedral shape and is not surface-treated.

3. ANALOGUE APPROACH JUSTIFICATION
The two analogue forms have the same structure. Under ambient atmosphere, the specific surface energy of particles increases with decreasing particle size. Therefore, particle aggregate to reach an energy minimum. The driving forces are hydrogen bonds and van der Waals forces (π-π interaction). Substantial energy is required to disperse the PR254 nanoform aggregates to particles that fall under the nanoform definition.
PR254 was been tested extensively addressing information requirements of Annexes VII to IX without identifying any biological target. PR254 nanoform could potentially have biological targets due to the different particle size distribution, which would require processes capable of dispersing the aggregates, e.g. in aqueous milieu. However, both forms have been tested according to OECD Test Guideline 318, demonstrating that PR254 nanoform cannot be dispersed under the condition of the study, i.e. immediately after sonification re-forms aggregates. Also, PR254 aggregates to a large extent, but can be more easily dispersed than the nanoform. The experiments demonstrated that exposure in aqueous milieu will be primarily to aggregates, regardless of the PR254 form.
Therefore, it is concluded that both forms will behave identically in studies, in which they are applied under atmospheric conditions and/or in aqueous milieus, so that for the PR254 nano-form no specific biological targets need to be considered.
As both forms form non-dispersible aggregates in aqueous milieu and under ambient conditions, read-across of toxicological studies from the source to the target form is scientifically justified.

4. DATA MATRIX
The data matrix is included as Annex 1 in the assessment report ‘PR254 bulk nano analogue approach 210111’ attached here below under ‘Attached justification’.
Reason / purpose for cross-reference:
reference to same study
Qualifier:
according to guideline
Guideline:
OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)
GLP compliance:
yes (incl. QA statement)
Type of study:
mouse local lymph node assay (LLNA)
Specific details on test material used for the study:
SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: A01297 (Cinilex DPP RED SR1C)
- Expiration date of the lot/batch: -
- Purity test date: -

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: room tempreature in the dark

Species:
mouse
Strain:
CBA
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Harlan UK Limited, Bicester, Oxon, UK
- Housing: individually
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19 -25 °C
- Humidity (%): 30-70%
- Air changes (per hr): 15/h
- Photoperiod (hrs dark / hrs light): 12/12

Vehicle:
acetone/olive oil (4:1 v/v)
Concentration:
5%, 10% or 25% w/w in acetone/olive oil 4:1
No. of animals per dose:
4
Key result
Parameter:
SI
Value:
0.99
Test group / Remarks:
5%
Parameter:
SI
Test group / Remarks:
10%
Remarks on result:
other: no results due to accidential spillage of pooled lymph node cell suspension
Key result
Parameter:
SI
Value:
1.13
Test group / Remarks:
25%
Parameter:
other: disintegrations per minute (DPM)
Remarks on result:
other: vehicle: 7694.44 5%: 7593.42 25% 8718.31

Due to accidental spillage of the single cell suspension of pooled lymph node cells for the group treated with the test material at a concentration of 10%, the results obtained cannot be verified as being accurate. For this reason the results for the 10% group were not used for reporting purposes. This was considered not to affect the purpose or integrity of the study.

Interpretation of results:
GHS criteria not met
Conclusions:
In a GLP-study according to OECD test guideline 429 (LLNA), the substance was not skin sensitising.
Executive summary:

A study was performed to assess the skin sensitisation potential of the test material in the CBA/Ca strain mouse following topical application to the dorsal surface of the ear.

Following a preliminary screening test, three groups, each of four animals, were treated with 50 p.1 (25 p1 per ear) of the test material as a suspension in acetone/olive oil 4:1 at concentrations of 5%, 10% or 25% w/w. A further group of four animals was treated with acetone/olive oil 4:1 alone.

Due to accidental spillage of the single cell suspension of pooled lymph node cells for the group treated with the test material at a concentration of 10%, the results obtained cannot be verified as being accurate. For this reason the results for the 10% group were not used for reporting purposes. This was considered not to affect the purpose or integrity of the study.

Simulation Indexes for 5% and 25 % were 0.99 and 1.13, respectively. Therefore, the test material was considered to be a non-sensitiser under the conditions of the test.

Endpoint:
skin sensitisation: in vitro
Data waiving:
study scientifically not necessary / other information available
Justification for data waiving:
other:
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (not sensitising)
Additional information:

The results of the LLNA is supported by two guinea pig maximisation tests, in which no indication of skin sensitisation were observed for the substances dosed as 0.3% and 3%, respectively.

Respiratory sensitisation

Endpoint conclusion
Endpoint conclusion:
no study available

Justification for classification or non-classification

Based on a reliable and GLP-compliant LLNA study, the substance is considered to be not a skin sensitiser. Therefore, no classification is warranted.