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Toxicological information

Skin sensitisation

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Administrative data

Endpoint:
skin sensitisation: in vivo (LLNA)
Type of information:
experimental study
Adequacy of study:
key study
Study period:
6 JULY 2018 to 16 APRIL 2019
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Justification for type of information:
TESTING PROPOSAL ON VERTEBRATE ANIMALS
[Please provide information for all of the points below. The information should be specific to the endpoint for which testing is proposed. Note that for testing proposals addressing testing on vertebrate animals under the REACH Regulation this document will be published on the ECHA website along with the third party consultation on the testing proposal(s).]

NON-CONFIDENTIAL NAME OF SUBSTANCE:
- Name of the substance on which testing is proposed to be carried out 1,4-bis(isopropylamino)anthraquinone

CONSIDERATIONS THAT THE GENERAL ADAPTATION POSSIBILITIES OF ANNEX XI OF THE REACH REGULATION ARE NOT ADEQUATE TO GENERATE THE NECESSARY INFORMATION [please address all points below]:
- Available GLP studies: OECD 442B
- Available non-GLP studies
- Historical human data
- (Q)SAR
- In vitro methods
- Weight of evidence: Log POW of test item >3.5. The OECD 442E shall not be considered by this reason.
- Grouping and read-across
- Substance-tailored exposure driven testing [if applicable]
- Approaches in addition to above [if applicable]
- Other reasons [if applicable]

CONSIDERATIONS THAT THE SPECIFIC ADAPTATION POSSIBILITIES OF ANNEXES VI TO X (AND COLUMN 2 THEREOF) OF THE REACH REGULATION ARE NOT ADEQUATE TO GENERATE THE NECESSARY INFORMATION:
- [free text]

FURTHER INFORMATION ON TESTING PROPOSAL IN ADDITION TO INFORMATION PROVIDED IN THE MATERIALS AND METHODS SECTION:
- Details on study design / methodology proposed [if relevant]

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2019
Report Date:
2019

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 442B (Skin Sensitization: Local Lymph Node Assay: BrdU-ELISA)
Version / remarks:
OECD Guideline No.442-B adopted 25 June 2018
Deviations:
no
GLP compliance:
yes (incl. certificate)
Type of study:
mouse local lymph node assay (LLNA): BrdU-ELISA

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
solid: particulate/powder

In vivo test system

Test animals

Species:
mouse
Strain:
CBA:J
Remarks:
CBA:JRj
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Elevage Janvier Labs (F-53941 Le Genest Saint Isle)
- Females (if applicable) nulliparous and non-pregnant: yes
- Microbiological status of animals, when known: not to contain any contaminant of a level that might have affected the purpose or integrity of the study.
- Age at study initiation: 8 weeks old
- Weight at study initiation: eman 20.6g
- Housing:The animals were individually housed in suspended solid-floor polypropylene cages furnished with softwood woodflakes.
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum):ad libitum
- Acclimation period: at least five days under stabling and nutritional conditions identical
to those of the test
- Indication of any skin lesions: none

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19°C to 25°C
- Humidity (%): 30% to 70%
- Air changes (per hr): The rate of air exchange was at least ten changes per hour
- Photoperiod (hrs dark / hrs light): lighting was controlled by a time switch to give twelve hours continuous light (07.00 to 19.00) and twelve hours darkness.
- IN-LIFE DATES: From: 29 AUGUST 2018 To: 3 SEPTEMBER 2018

Study design: in vivo (LLNA)

Vehicle:
dimethylformamide
Concentration:
50%, 25%, 10%
No. of animals per dose:
4
Details on study design:
PRE-SCREEN TESTS:
- Compound solubility: 50%
- Irritation: no signs
- Systemic toxicity: no signs of excessive irritation
- Ear thickness measurements: 3 (Day 1: 0.20, Day 3: 0.20, Day 6: 0.22)
- Erythema scores: no signs

MAIN STUDY

ANIMAL ASSIGNMENT AND TREATMENT
- Name of test method: ELISA TEST
- Criteria used to consider a positive response:
% increase in ear thickness between day 1 and day 3 and/or between day 1 and day 6
<10% --> non irritant
10-25% --> slightly irritant
>25% --> irritant

TREATMENT PREPARATION AND ADMINISTRATION:
The mice were treated by daily application of 25 µL of the appropriate concentration of the test item to the dorsal surface of each ear for three consecutive days (Days 1, 2 and 3). The test item formulation was administered using an automatic micropipette and spread over the dorsal surface of the ear using the tip of the pipette.
Positive control substance(s):
hexyl cinnamic aldehyde (CAS No 101-86-0)
Statistics:
SI: Stimulation Index.

Results and discussion

In vivo (LLNA)

Resultsopen allclose all
Key result
Parameter:
SI
Value:
ca. 0.94
Variability:
SD 0.17
Test group / Remarks:
Dose 10%
Mean of 1 run with 4 replicates, each SI<1.6
Remarks on result:
not determinable
Key result
Parameter:
SI
Value:
ca. 0.93
Variability:
SD 0.24
Test group / Remarks:
Dose 25%
mean of 1 run with 4 replicates, each SI <1.6
Remarks on result:
not determinable
Key result
Parameter:
SI
Value:
ca. 0.8
Variability:
SD 0.12
Test group / Remarks:
Dose 50%
Mean of 1 run with 4 replicates each SI <1.6.
Remarks on result:
not determinable
Cellular proliferation data / Observations:
CELLULAR PROLIFERATION DATA

DETAILS ON STIMULATION INDEX CALCULATION

EC3 CALCULATION

CLINICAL OBSERVATIONS:
Blue coloration not preventing quotation of erythema was noted in all animals treated at 10%, 25%
and 50%, between days 2 and 6.
No increase in ear thickness and in ear weight was noted in animals treated at 10%, 25% and 50%.
Therefore, the test item has to be considered as not excessively irritant at these concentrations.

BODY WEIGHTS
The mean of weights in each run, was that the animals gained weight.

Any other information on results incl. tables

TEST ITEM: 1,4-bis(isopropylamino) anthraquinone
Date of 1st application (day 1): 29 August 2018 Vehicle: N,N-dimethylformamide
Table 3








































































































































































Body weight gain
(g)
Bodyweight (g)Groups Test itemAnimals No.
Day 1 Day 6
Sf 1001 20.0 20.4 0.4DMF1
Sf 1002 21.9 21.9 0.0
Sf 1003 21.3 22.3 1.0
Sf 1004 19.0 20.4 1.4
20.6 21.3 0.7MEAN
1.3 1.0 0.6Standard-deviation
Sf 1036 21.0 21.6 0.610%2
Sf 1037 19.3 19.9 0.6
Sf 1038 20.7 20.5 -0.2
Sf 1039 23.6 24.3 0.7
21.2 21.6 0.4MEAN
1.8 1.9 0.4Standard-deviation
Sf 1041 21.9 23.2 1.3325%
Sf 1042 19.7 24.1 4.4
Sf 1043 21.2 21.8 0.6
Sf 1044 19.2 19.9 0.7
20.5 22.3 1.8MEAN
1.3 1.8 1.8Standard-deviation
Sf 1046 20.6 20.7 0.1450%
Sf 1047 20.5 21.3 0.8
Sf 1048 20.5 20.5 0.0
Sf 1050 20.7 21.6 0.9
20.6 21.0 0.5MEAN
0.1 0.5 0.5Standard-deviation

DMF: N,N-dimethylformamide

Applicant's summary and conclusion

Interpretation of results:
GHS criteria not met
Conclusions:
The results obtained, under these experimental conditions, enable to conclude that the test item
1,4-bis(isopropylamino) anthraquinone does not have to be classified as a skin sensitizer, in
accordance with the Regulation EC No. No. 1272/2008 on classification, labelling and packaging of substances and mixtures.
No signal word or hazard statement is required.