N-(2,3-dihydro-2-oxo-1H-benzimidazol-5-yl)-2-[(2-methoxyphenyl)azo]-3-oxobutyramide

Endpoint summary

Description of key information

In the a LLNA (OECD 429) the test item suspended in ethanol:water (7 +3) was assessed for its possible contact allergenic potential.

For this purpose a local lymph node assay was performed using test item concentrations of 2.5, 5 and 10%.

The animals did not show any clinical signs during the course of the study and no cases of mortality were observed. Due to the yellow colour of the test item local irritation reactions such as ear redness could not be examined.

In this study Stimulation Indices (S.I.) of 1.48, 1.51 and 1.63 were determined with the test item at concentrations of 2.5, 5 and 10% (w/v) in ethanol:water (7+3), respectively.

The test item was not a skin sensitiser in this LLNA-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 vivo (LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

From 4 APR 2006 to 25 APR 2006

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)

Deviations:

no

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: 6-12 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 (%): 27-70
- Photoperiod (hrs dark / hrs light): 12/12

Vehicle:

other: ethanol:water (7+3)

Concentration:

0, 2.5, 5, and 10% (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: ethanol:water (7+3) was selected because the test item suspended best in this vehicle
- 10% (w/v) was the highest technically applicable concentration
- Irritation: no severe local irritation or systemic toxicity after single application of test item concentrations of 2.5, 5 or 10% (w/v); irritating effects such as redness could not be detected due to yellow staining at concentrations of 2.5, 5 and 10% (w/v)
- 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

Positive control results:

Stimulation indices of 6.08, 13.59 and 26.41 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 could not be calculated, since all SI's are above 3.

Parameter:

SI

Remarks on result:

other: Stimulation indices (SI) were all below 3. The following SI were calculated: 2.5% test item: 1.48 5% test item: 1.51 10% test item: 1.63

Parameter:

other: disintegrations per minute (DPM)

Remarks on result:

other: Background of measurement: 41.88 - 106.47; mean: 74.18 (was taken for calculations) control group: 2728.98 2.5% test item: 3996.75 5% test item: 4091.9 10% test item: 4389.52

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)	Group	Measurement DPM	Calculation			Result
			DPM-BGa)	number of lymph nodes	DPM per lymph node^b)	S.I.
---	BG I	41.88	---	---	---	---
---	BG II	106.47	---	---	---	---
---	CG 1	2728.98	2654.8	8	331.9
2.5	TG 2	3996.75	3922.6	8	490.3	1.48
5	TG 3	4091.90	4017.7	8	502.2	1.51
10	TG 4	4389.52	4315.3	8	539.4	1.63

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:

not sensitising

Remarks:

Migrated information Criteria used for interpretation of results: EU

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 suspended in ethanol:water (7 +3) was assessed for its possible contact allergenic potential.

For this purpose a local lymph node assay was performed using test item concentrations of 2.5, 5 and 10%.

The animals did not show any clinical signs during the course of the study and no cases of mortality were observed. Due to the yellow colour of the test item local irritation reactions such as ear redness could not be examined.

In this study Stimulation Indices (S.I.) of 1.48, 1.51 and 1.63 were determined with the test item at concentrations of 2.5, 5 and 10% (w/v) in ethanol:water (7+3), respectively.

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

The test material did not show adverse effects in a local lymph node assay in mice.