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Diss Factsheets

Toxicological information

Skin sensitisation

Currently viewing:

Administrative data

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
Remarks:
performed in accordance with OECD and GLP guidelines

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2006
Report date:
2005

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Remarks:
according to German Chemikaliengesetzt and OECD Principles of Good Laboratory Practice
Type of study:
mouse local lymph node assay (LLNA)

Test material

Constituent 1
Chemical structure
Reference substance name:
2,20-dichloro-13,31-diethyl-4,22-dioxa-13,18,31,36-tetraazanonacyclo[19.15.0.0³,¹⁹.0⁵,¹⁷.0⁶,¹⁴.0⁷,¹².0²³,³⁵.0²⁴,³².0²⁵,³⁰]hexatriaconta-1(36),2,5,7(12),8,10,14,16,18,20,23(35),24(32),25,27,29,33-hexadecaene
EC Number:
606-790-9
Cas Number:
215247-95-3
Molecular formula:
C34H22Cl2N4O2
IUPAC Name:
2,20-dichloro-13,31-diethyl-4,22-dioxa-13,18,31,36-tetraazanonacyclo[19.15.0.0³,¹⁹.0⁵,¹⁷.0⁶,¹⁴.0⁷,¹².0²³,³⁵.0²⁴,³².0²⁵,³⁰]hexatriaconta-1(36),2,5,7(12),8,10,14,16,18,20,23(35),24(32),25,27,29,33-hexadecaene
Test material form:
solid: nanoform

In vivo test system

Test animals

Species:
mouse
Strain:
CBA
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
-strain: CBA/CaOlaHsd
- Source: Harlan Netherlands
- Age at study initiation: 8-12 weeks (beginning of acclimatization)
- Weight at study initiation: 18.3 g to 21.4 g
- Housing: single caging, Makrolon Type I, with wire mesh top, granulated soft wood bedding
- Diet (e.g. ad libitum): Pelleted standard diet, ad libitum
- Water (e.g. ad libitum): tap water, ad libitum
- Acclimation period: yes

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 +/- 3°C
- Humidity (%): 30 - 70 %
- Air changes (per hr): no data
- Photoperiod (hrs dark / hrs light): artificial light 6.00 a.m. - 6.00 p.m.

Study design: in vivo (LLNA)

Vehicle:
acetone/olive oil (4:1 v/v)
Concentration:
0 (control), 7.5, 15 and 30% (w/v)
No. of animals per dose:
4 animals per treatment group, 4 animals in the control group
Details on study design:
RANGE FINDING TESTS:
- Compound solubility:
To determine the highest non-irritant test concentration or the highest technically
applicable concentration, a pretest was performed under RCC-CCR non-GLP study
conditions in two mice. The data showed that the highest test item concentration,
which could be technically used was a 30 % suspension in acetone:olive oil, 4:1 (v/v).
Also in other vehicles used, e.g DMSO, DMF a higher concentrations could not
be suspended.
- Irritation:
At this concentration the treated mouse did not show any toxic symptoms. Due to the colour of
the test item irritation reactions of the ear skin such as redness could not be observed. No
swelling of the ears was observed.
- Lymph node proliferation response: not evaluated


MAIN STUDY
ANIMAL ASSIGNMENT AND TREATMENT
- Name of test method: Pooled (4 animals/group) LLNA
- Criteria used to consider a positive response:
A test item is regarded as a sensitiser in the LLNA if the following criteria are fulfilled:
- First, that exposure to at least one concentration of the test item resulted in an
incorporation of 3HTdR at least 3-fold or greater than that recorded in control mice, as
indicated by the stimulation index.
- Second, that the data are compatible with a conventional dose response, although
allowance must be made (especially at high topical concentrations) for either local
toxicity or immunological suppression.


TREATMENT PREPARATION AND ADMINISTRATION:
Four female mice were treated with different concentrations of the test item and vehicle alone by topical application at the dorsum of each ear lobe (left and right) on three consecutive days. Five days after the first topical application, the mice were intravenously injected into a tail vein with radio-labelled thymidine (3H-methyl thymidine). Five hours after intravenous injection, the mice were sacrificed and the draining auricular lymph nodes excised and pooled per group. Single cell suspensions of lymph node cells were prepared from pooled lymph nodes which were subsequently washed and incubated with trichloroacetic acid overnight. The proliferative capacity of pooled lymph node cells was determined by the incorporation of 3H-methyl thymidine measured in a ß-scintillation counter.
Positive control substance(s):
hexyl cinnamic aldehyde (CAS No 101-86-0)
Statistics:
-

Results and discussion

Positive control results:

Test item concentration %(w/v) Group Measurement DPM DPM-BG a) Number of lymph nodes DPM per lymph node b) S.I.
- BG I 0.0 - - - -
- BG II 1.2 - - - -
- CG 1 3855.2 3854.6 8 481.8
5 TG 1 7544.2 7543.6 8 943.0 2.0
10 TG 2 11686.7 11686.1 8 1460.8 3.0
25 TG 3 18946.4 18945.8 8 2368.2 4.9
BG = Background (1 ml 5% trichloroacetic acid) in duplicate
CG = Control Group (Vehicle: acetone:olive oil (4+1))
TG = Test Group
S.I. = Stimulation Index
a) = The mean value was taken from the figures BG I and BG II
b) = Since the lymph nodes of the animals of a dose group were pooled, DPM/node was determined by dividing the measured value by the number of lymph nodes pooled

EC3 = (a-c) [(3-d)/(b-d)] + c = 9.9%(w/v)

In vivo (LLNA)

Resultsopen allclose all
Key result
Parameter:
SI
Value:
0.6
Test group / Remarks:
7.5 %
Key result
Parameter:
SI
Value:
0.7
Test group / Remarks:
15 %
Key result
Parameter:
SI
Value:
0.6
Test group / Remarks:
30 %
Key result
Parameter:
EC3
Test group / Remarks:
all
Remarks on result:
other: The EC3 value could not be calculated, since none of the tested concentrations induced an S.I. greater than 3.

Any other information on results incl. tables

Test item concentration% (w/v)   Measurement dpm     Calculation      Result
    dpm - BG a) number of lymph nodes dpm per lymph node b)  S.I. 
 -- BG I  4.8  -- -- -- --
 -- BG II   0.0  -- -- --  -- 
 --  CG 1  8271.0  8268.6  8 1033.6  --
7.5  TG 1  4362.4  4360.0  8 545.0 0.5
15  TG 2  4287.0 4284.6  8 714.1 0.7
30  TG 3  5001.7  4999.3  8 624.9 0.6

BG = Background (1 ml 5% trichloroacetic acid) in duplicate

CG = Control Group

TG = Test Group

S.I. = Stimulation Index

a) = The mean value was taken from the figures BG I and BG II

b) = Since the lymph nodes of the animals of a dose group were pooled, DPM/Node was determined by dividing the measured value by the number of lymph nodes pooled

The EC3 value could not be calculated, since all SI's are below.

No deaths occurred during the study period.

Due to the intense violett colour of the test item local irritation reactions such as ear redness could not be detected. However, no swelling of the ears was observed. Body weights were within the normal range.

Applicant's summary and conclusion

Interpretation of results:
GHS criteria not met
Conclusions:
No classification
The test item (Pigment Violet 23) was found to be not a skin sensitiser in the LLNA when tested at up to 30% in acetone:olive oil 4:1 (v/v).
Executive summary:

In the study the test item dissolved in acetone:olive oil, 4:l (v/v) was assessed for its possible contact allergenic potential. For this purpose a local lymph node assay was performed using test item concentrations of 7.5, 15, 30 %. The animals did not show any clinical signs during the course of the study and no cases of mortality were observed. In this study Stimulation indices (S.I.) of 0.5, 0.7, and 0.6 were determined with the test item at concentrations of 7.5, 15, 30 %(w/v) in acetone:olive oil, 4:l (v/v). The test item was found to be not a skin sensitiser in this assay.