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Diss Factsheets

Administrative data

Endpoint:
skin sensitisation: in vivo (LLNA)
Type of information:
experimental study
Adequacy of study:
key study
Study period:
from 13AUG2009 to 17SEP2009
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP compliance statement. Follows outlined guidelines

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2009
Report date:
2009

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to guideline
Guideline:
OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)
Qualifier:
according to guideline
Guideline:
EU Method B.42 (Skin Sensitisation: Local Lymph Node Assay)
Qualifier:
according to guideline
Guideline:
EPA OPPTS 870.2600 (Skin Sensitisation)
GLP compliance:
yes
Type of study:
mouse local lymph node assay (LLNA)

Test material

Constituent 1
Reference substance name:
Adiponitrile
EC Number:
203-896-3
EC Name:
Adiponitrile
Cas Number:
111-69-3
IUPAC Name:
adiponitrile
Details on test material:
Adiponitrile >99% purity

In vivo test system

Test animals

Species:
mouse
Strain:
CBA
Sex:
female
Details on test animals and environmental conditions:
Mouse, CBA/J strain, inbred, SPF-Quality. 20 females (nulliparous and non-pregnant), five females per group. Young adult animals (approx. 11 weeks old) were selected. Body weight variation was within +/- 20% of the sex mean. A health inspection was performed prior to treatment, to ensure that the animals are in a good state of health. Special attention was paid to the ears, which were intact and free from any abnormality. The results of a reliability test with Hexylcinnamaldehyde, performed not more than 6 months previously, are summarized in the appendix. Similar procedures were used in the reliability test and in this study.
Conditions
Animals were housed in a controlled environment, in which optimal conditions were considered to be approximately 15 air changes per hour, a temperature of 21.0 ± 3.0ºC (actual range: 21.6 - 23.5ºC), a relative humidity of 40-70% (actual range: 42 - 74%) and 12 hours artificial fluorescent light and 12 hours darkness per day.
Accommodation
Individual housing in labeled Macrolon cages (MI type; height 12.5 cm) containing sterilized sawdust as bedding material (Litalabo, S.P.P.S., Argenteuil, France). Paper (Enviro-dri, Wm. Lillico & Son (Wonham Mill Ltd), Surrey, United Kingdom) was supplied as cage-enrichment.
Acclimatization period
The acclimatization period was at least 5 days before the start of treatment under laboratory conditions. Accommodation was as described above except that the animals were group housed in Macrolon cages (MIII type; height 18 cm).
Diet
Free access to pelleted rodent diet (SM R/M-Z from SSNIFF® Spezialdiäten GmbH, Soest, Germany).
Water
Free access to tap water.
Results of analysis for diet (nutrients and contaminants), sawdust, paper and water were assessed and did not reveal any findings that were considered to have affected the study integrity. All certificates and results of analysis are retained in the NOTOX archives.

Study design: in vivo (LLNA)

Vehicle:
dimethylformamide
Concentration:
induction (test substance; % w/w)
0 (Dimethyl formamide)
25
50
100
No. of animals per dose:
5
Details on study design:
Skin reactions / Irritation
No irritation or oedema of the ears was observed in any of the animals examined.
Macroscopy of the auricular lymph nodes and surrounding area (Table 2)
All auricular lymph nodes of the animals of the experimental and control groups were considered normal in size, except for one node of one control animal which was considered small in size.
Body weights
Body weights and body weight gain of experimental animals remained in the same range as controls over the study period. The slight body weight loss, noted in some animals, was considered not toxicologically significant.
Radioactivity measurements
Mean DPM/animal values for the experimental groups treated with test substance concentrations 25, 50 and 100% were 314, 448 and 610 respectively.
The mean DPM/animal value for the vehicle control group was 443.
Toxicity and mortality
One animal (at 100%) was found dead and one moribund animal (at 50%) was sacrificed for ethical reasons on Day 2 prior to dosing. The moribund animal showed hunched and flat posture, piloerection, laboured breathing, ptosis and hypothermia on Day 2.
Since no further mortality occurred and no symptoms of systemic toxicity were observed in the surviving animals of the main study, these deaths were considered to be of incidental nature and not test substance related.
Positive control substance(s):
hexyl cinnamic aldehyde (CAS No 101-86-0)
Statistics:
A reliability check is carried out at regular intervals to check the sensitivity of the test system and the reliability of the experimental techniques as used by NOTOX. In this study, performed in February 2009, females of the CBA/J mouse strain (from Charles River France, L¿Arbresle Cedex, France) were checked for the sensitivity to Alpha- Hexylcinnamaldehyde, technical grade. The females were approx. 10 weeks old at commencement of the study. The study was based on the OECD Guideline No. 429, EC No 440/2008, Part B.42 and EPA, OPPTS 870.2600 ¿Skin Sensitization¿. Alpha-hexylcinnamicaldehyde, tech. 85% (CAS no. 101-86-0) was fabricated under lot no. 13102MO (Sigma- Aldrich, Steinheim, Germany). Concentrations used for this study were 5, 10 and 25% in Acetone/Olive oil (4:1 (v/v)).

Results and discussion

Positive control results:
The SI values calculated for the substance concentrations 5, 10 and 25% were 2.0, 2.2 and 4.2 respectively. An EC3 value of 16.0 % was calculated using linear interpolation. The calculated EC3 value was found to be in the acceptable range of 2 and 20%. The results of the 6 monthly HCA reliability checks of the recent years were 13.1, 15.6, 14.1, 13.8 and 13.9%.
Based on the results, it was concluded that the Local Lymph Node Assay as performed at NOTOX is an appropriate model for testing for contact hypersensitivity.
The raw data, protocol and report from this study are kept in the NOTOX archives. The test described above was performed in accordance with NOTOX Standard Operating Procedures and the report was audited by the QA-unit.

In vivo (LLNA)

Resultsopen allclose all
Parameter:
SI
Remarks on result:
other: group 2 0.7 +/- 0.2 group 3 1.0 +/- 0.2 group 4 1.4 +/- 0.4 group 1 1. +/- 0.3
Parameter:
other: disintegrations per minute (DPM)
Remarks on result:
other: see Remark
Remarks:
1-399 2-342 3-448 4-779 5-247 6-243 7-244 8-211 9-415 10-457 11-544 12-423 13-454 14-371 15-Animals were found dead or sacrificed for ethical reasons on Day 2 16-Animals were found dead or sacrificed for ethical reasons on Day 2 17-825 18-707 19-394 20-513 group 2 mean 314 +/- 51 group 3 mean 448 +/- 36 group 4 mean 610 +/- 96 group 1 mean 443 +/- 90

Any other information on results incl. tables

 group  test substance (%w/w)  mean DPM +/- SEM  SI +/- SEM
 2  25  314 ± 51  0.7 ± 0.2
 3  50  448 ± 36  1.0 ± 0.2
 4  100  610 ± 96  1.4 ± 0.4
 1  0 (vehicle)  443 ± 90  1.0 ± 0.3

Applicant's summary and conclusion

Interpretation of results:
not sensitising
Remarks:
Migrated information
Conclusions:
ADIPONITRILE (ADN) would not be regarded as skin sensitizer and has no obligatory labeling requirement for sensitization by skin contact according to the Globally Harmonized System of Classification and Labeling of Chemicals (GHS) of the United Nations (2007) and the Regulation (EC) No 1272/2008 on classification, labeling and packaging of substances and mixtures.
Executive summary:

According to an OECD 429 study, the test substance did not ellicit an SI value of >=3 on 100% adiponitrile and therefore ADN would be considered a non-sensitizer.

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