2,2'-[ethylenebis(oxyphenyl-2,1-eneazo)]bis[N-(2,3-dihydro-2-oxo-1H-benzimidazol-5-yl)-3-oxobutyramide

Endpoint summary

Description of key information

In the study Pigment Yellow 180, dissolved in DMF, was assessed for its possible contact allergenic potential.

For this purpose a local lymph node assay was performed using test item concentrations of 5.0 %, 10.0 %, and 15.0 %.

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 2.42, 2.77, and 2.28 were determined at concentrations of 5.0, 10.0, and 15.0 (w/v) in DMF.

The test item was not a skin sensitiserin this assay.

Key value for chemical safety assessment

Skin sensitisation

Link to relevant study records

Reference

Reference 1

Endpoint:

skin sensitisation: in vitro

Data waiving:

study scientifically not necessary / other information available

Justification for data waiving:

an in vitro skin sensitisation study does not need to be conducted because adequate data from an in vivo skin sensitisation study are available

Reference 2

Endpoint:

skin sensitisation: in chemico

Data waiving:

study scientifically not necessary / other information available

Justification for data waiving:

an in vitro skin sensitisation study does not need to be conducted because adequate data from an in vivo skin sensitisation study are available

Reference 3

Endpoint:

skin sensitisation: in vivo (LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

From 10 AUG 2004 to 1 DEC 2004

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Guideline study (OECD TG 429)

Qualifier:

according to guideline

Guideline:

OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)

GLP compliance:

yes

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 Netherlands BV, Horst, The Netherlands
- Age at study initiation: 8-12 weeks
- Housing: Individually in Macrolon Type I cages with wire mesh top and granulated sooft wood bedding
- Diet: Pelleted standard diet (provided by Harlan Winkelmann GmbH, Borchen, Germany); ad libitum
- Water: Community tap water from Rossdorf, ad libitum
- Acclimation period: yes, but duration not mentioned

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22+/-3
- Humidity (%): 30-74
- Photoperiod (hrs dark / hrs light): 12/12

Vehicle:

dimethylformamide

Concentration:

0, 5, 10, and 15% (w/v)

No. of animals per dose:

4 females per dose group
2 females in the pre-test

Details on study design:

RANGE FINDING TESTS:
- Compound solubility: DMF was selected as solvent, because with this vehicle the highest practically applicable test item concentration could be achieved
- 15% (w/v) was the highest technically applicable concentration whilst avoiding systemic toxicity and excessive local irritation
- Lymph node proliferation response: no data

MAIN STUDY
ANIMAL ASSIGNMENT AND TREATMENT
- Name of test method: LLNA
- Criteria used to consider a positive response:
1: exposure to at least one concentration of the test item resulted in an incorporation of 3H-methyl thymidine at least 3-fold or greater than that recorded in control mice as indicated by the stimulation index
2: data are compatible with a conventional dose response, although allowance must be made for either local toxicity or immunological suppression

TREATMENT PREPARATION AND ADMINISTRATION:
- individual preparation of weight volume dilutions using a magnetic stirrer as homogenizer
- test item preparations were made feshly before each dosing occasion
- 25 µl were spread over the entire dorsal surface of each ear lobe once daily for three consecutive days

Positive control substance(s):

hexyl cinnamic aldehyde (CAS No 101-86-0)

Statistics:

- calculations of mean values and standard deviations for body weight
- the EC3 value was calculated according to the equation EC3 = (a-c) [(3-d)/(b-d)] + c; where EC3 is the estimated concentration of the test item required to produce a 3-fold increase in draining Iymph node cell proliferative activity; (a, b) and (c, d) are respectively the co-ordinates of the two pair of data lying immediately above and below the value of 3 on the local lymph node assay dose response Plot.

Positive control results:

Stimulation indices of 1.9, 6.1 and 11.5 were determined with the positive control item at concentrations of 5%, 10% and 25% (w/v), respectively, in acetone:olive oil, 4:1 (v/v). An EC3 value was calculated (= 6.3% (w/v)).
Results of the second positive control test revealed stimulation indices of 2.0, 3.0 and 4.9 were determined with the positive control item at concentrations of 5%, 10% and 25% (w/v), respectively, in acetone:olive oil, 4:1 (v/v). An EC3 value was calculated (= 9.9% (w/v)).

Key result

Parameter:

SI

Value:

2.42

Test group / Remarks:

5%

Key result

Parameter:

SI

Value:

2.77

Test group / Remarks:

10%

Key result

Parameter:

SI

Value:

2.28

Test group / Remarks:

15%

Parameter:

other: disintegrations per minute (DPM)

Remarks on result:

other: Background of measurement: 0 - 1.2; mean: 0.6 (was taken for calculations) control group: 1851.6 5% test item: 4472.8 10% test item: 5128.8 15% test item: 4214.4

The animals did not show any clinical signs during the course of the study and no cases of mortality were observed. Calculation of the EC3 value was not performed because no test concentration produced a SI of 3 or higher.

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	0.0	---	---	---	---
---	BG II	1.2	---	---	---	---
---	CG 1	1851.6	1851.0	8	231.4
5.0	TG 2	4472.8	4472.2	8	559.0	2.42
10.0	TG 3	5128.8	5128.2	8	641.0	2.77
15.0	TG 4	4214.4	4213.8	8	526.7	2.28

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

Interpretation of results:

GHS criteria not met

Remarks:

Regulation (EC) No. 1272/2008

Conclusions:

Tested in this study according to OECD test guideline no. 429 and GLP conditions the test item was not a skin sensitizer.

Executive summary:

In the study the test item dissolved in DMF was assessed for its possible contact allergenic potential.

For this purpose a local lymph node assay was performed using test item concentrations of 5.0 %, 10.0 %, and 15.0 %.

The animals did not show any clinical signs during the course of the study and no cases of mortality observed.

In this study Stimulation Indeces (S.I.) of 2.42, 2.77, and 2.28 were determined with the test item at concentrations of 5.0, 10.0, and 15.0 (w/v) in DMF.

The test item was not a skin sensitiser in this assay.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not sensitising)

Respiratory sensitisation

Endpoint conclusion

Endpoint conclusion:

no study available

Justification for classification or non-classification

No classification: No adverse effects observed