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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
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2018

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 442B (Skin Sensitization: Local Lymph Node Assay: BrdU-ELISA)
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

In vivo test system

Test animals

Species:
mouse
Strain:
CBA:J
Sex:
female
Details on test animals and environmental conditions:
The animals were nulliparous and non-pregnant. After an acclimatisation period of at least five days under stabling and nutritional conditions identical to those of the test, the animals were selected at random and given a number unique within the study by indelible ink-marking on the tail and a number written on a cage card. At the start of the main study the animals were 8 or 9 weeks old. The animals were weighed at the beginning and at the end of the study.

Housing:
The animals were individually housed in suspended solid-floor polypropylene cages furnished with softwood woodflakes. The temperature and relative humidity were controlled to remain within target ranges of 19°C to 25°C and 30% to 70%, respectively. The rate of air exchange was at least ten changes per hour and the lighting was controlled by a time switch to give twelve hours continuous light (07.00 to 19.00) and twelve hours darkness. The animals were provided with environmental enrichment items which were considered not to contain any contaminant of a level that might have affected the purpose or integrity of the study.

Food and drink:
The drinking water and food were supplied ad libitum. Microbiological and chemical analyses of the water were carried out once every six months.

Study design: in vivo (LLNA)

Vehicle:
acetone/olive oil (4:1 v/v)
Concentration:
20%, 10%, 5% of test item diluted in acetone/olive oil
No. of animals per dose:
4 mice/dose group
Details on study design:
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). A further group of four mice received the vehicle alone in the same manner.

Clinical observations and mortality: All animals were observed daily on Days 1, 2, 3, 4, 5 and 6. Any signs of toxicity or signs of ill health during the test were recorded.
Body weights: The bodyweight of each mouse was recorded on Day 1 (prior to dosing) and Day 6 (prior to termination).
Ear thickness measurements and recording of local reactions: Ear thickness measurements and recording of local reactions were performed in order to assess any possible irritant effect of the test item, as possible irritancy may be involved in false positive lymphoproliferative responses. On day 1 and on day 3 (before application) as well as on day 6 (after sacrifice) of each experiment, the thickness of the right ear of each animal of the vehicle control and treated groups was measured by a micrometer. Furthermore, on day 6, punch biopsies of 8 mm in diameter of the apical area of both ears were prepared and weighed in order to assess the irritation potential of the test item and the two lymph nodes per mouse were weighed. Any irritation reaction (erythema and oedema) was recorded in parallel. Any other observation (dryness, presence of residual test item…) was noted.
Positive control substance(s):
hexyl cinnamic aldehyde (CAS No 101-86-0)

Results and discussion

Positive control results:
The substance a-Hexylcinnamaldehyde in accordance with the Regulation (EC) No. 1272/2008 has to be classified in category 1 “Skin sensitisation”, based on the values of EC1.6.

In vivo (LLNA)

Resultsopen allclose all
Key result
Parameter:
SI
Value:
ca. 1.09
Test group / Remarks:
5% dose group
Key result
Parameter:
SI
Value:
ca. 0.91
Test group / Remarks:
10% dose group
Key result
Parameter:
SI
Value:
ca. 0.88
Test group / Remarks:
20% dose group

Any other information on results incl. tables

Clinical observations and mortality: no mortality and no signs of systemic toxicity were noted in the test and control animals during the test.

Weight evolution: bodyweight changes of the test animals between Day 1 and Day 6 were comparable to those observed in the corresponding control group animals over the same period.

Estimation of the proliferative response of lymph node cells: the Stimulation Index (SI) calculated by individual approach was 1.09, 0.91 and 0.88 for the treated groups at 5%, 10% and 20%, respectively.

Therefore, the EC1.6 cannot be determined due to the absence of SI value higher than 1.6.

Local irritation: slight dryness of the skin was noted in animals treated at 10% and 20% on day 6. A slight increase in ear weight (+9.5%, +10.3% and +6.1%) was noted in animals treated at 5%, 10%

and 20% respectively. Therefore, the test item has to be considered as not excessively irritant at these concentrations, in accordance with the O.E.C.D. criteria.

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 Gamma Cobalt 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.