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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
Study period:
25 April 2012 to 14 May 2012
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Study conducted under GLP conditions

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2012
Report Date:
2012

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)
Deviations:
yes
Remarks:
*Temp. dev. from the min. level of daily mean rel. humidity:Hist.data don’t indicate effect *Subst wasn’t flushed with N2:subst was weighted,kept in closed cont. & used within 2 h following weighing.Don’t expect that H2O content of air affected integrity.
Qualifier:
according to
Guideline:
EU Method B.42 (Skin Sensitisation: Local Lymph Node Assay)
Deviations:
yes
Remarks:
*Temp. dev. from the min. level of daily mean rel. humidity:Hist.data don’t indicate effect *Subst wasn’t flushed with N2:subst was weighted,kept in closed cont. & used within 2 h following weighing.Don’t expect that H2O content of air affected integrity.
Qualifier:
according to
Guideline:
EPA OPPTS 870.2600 (Skin Sensitisation)
Deviations:
yes
Remarks:
*Temp. dev. from the min. level of daily mean rel. humidity:Hist.data don’t indicate effect *Subst wasn’t flushed with N2:subst was weighted,kept in closed cont. & used within 2 h following weighing.Don’t expect that H2O content of air affected integrity.
GLP compliance:
yes
Type of study:
mouse local lymph node assay (LLNA)

Test material

Reference
Name:
Unnamed
Type:
Constituent
Type:
Constituent
Test material form:
other: Liquid
Details on test material:
Name of test material (as cited in study report: MTDID 20422
- Substance type: Pure active substance
- Physical state: Liquid
- Analytical purity: 99.99%
- Purity test date: 02/16/2011
- Expiration date of the lot/batch: 02/16/2013
- Storage condition of test material: At room temperature in the dark.

In vivo test system

Test animals

Species:
mouse
Strain:
other: CBA/J Strain, inbred, SPF-Quality
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Janvier, Le Genest-Saint-Isle, France
- Age at study initiation: Approx. 10 weeks old
- Weight at study initiation: 20-24 grams
- Housing: Labeled Makrolon cages (MIII type; height 18cm) containing sterized sawdust as bedding material (Litalabo, S.P.P.S., Argenteuil, France)
- Diet (e.g. ad libitum): Pelleted rodent diet (SM R/M-Z from SSNIFF Spezialdaten GmbH, Soest, Germany) ad libitum.
- Water (e.g. ad libitum): Tap water ad libitum.
- Acclimation period: At least 5 days before start of treatment.
ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18-24
- Humidity (%): 40-70
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 12/12
IN-LIFE DATES: From: 25 April 2012To: 14 May 2012

Study design: in vivo (LLNA)

Vehicle:
other: Water (Elix, Millipore S.A.S., Molsheim, France) with 1% pluronic L92 (BASF, New Jersey, USA)
Concentration:
0, 25, 50, and 100 % (w/w) of the test subtance
No. of animals per dose:
5
Details on study design:
RANGE FINDING TESTS:
- Compound solubility: Reacts with water, the hydrate is soluble up to 75%
- Irritation: Very slight erythema of the ears was observed at 100% test substance (w/w)
- Lymph node proliferation response:
MAIN STUDY
ANIMAL ASSIGNMENT AND TREATMENT
- Name of test method:
- Criteria used to consider a positive response: An SI value greater than or equal to 3 signifies that the test substance may be regarded as a skin sensitizer.
TREATMENT PREPARATION AND ADMINISTRATION
The dorsal suface of both ears was topically treated (25 microliters/ear) with the test substance concentration. Dosing was done using directed air extraction equipment.
Positive control substance(s):
hexyl cinnamic aldehyde (CAS No 101-86-0)

Results and discussion

In vivo (LLNA)

Resultsopen allclose all
Parameter:
SI
Remarks on result:
other: 25%: 1.4 50%: 2.3 100%: 0.8
Parameter:
other: disintegrations per minute (DPM)
Remarks on result:
other: 25%: 603 50%: 1004 100%: 370

Applicant's summary and conclusion

Interpretation of results:
not sensitising
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
The SI values for the 25, 50, and 100% concentrations were 1.4, 2.3, and 0.8 respectively. No EC3 value was calculated because all SI values were <3.There was very slight erythema of the ears in animals treated with the 50% and 100% concentrations of the test material, but was considered not toxicologically significant. No mortality, clinical signs of toxicity, or significant body weight change were observed. All auricular lymph nodes were considered normal in size. Based on the results of the test, extracts of the test article showed no evidence of dermal sensitization.
Executive summary:

The dermal sensitization potential of the test article (clear colorless liquid, purity 99.99%) was evaluated in the local lymph node assay (LLNA) with CBA/J strain mice. This study was performed in compliance with OECD GLP ENV/MC/CHEM (98)17 (1997). The test method was based on OECD Section 4 No. 429 (2010), EC No. 440/2008 B42, and EPA OPPTS 870.2600 (2003). The test article was prepared in 1% watery pluronic L92 (as approved by the sponsor). A pre-screen test was conducted to select the highest test article concentrations of 100% and 50%. Female mice (5/treatment) received the control (1% L92 in water), 25%, 50%, or 100% concentrations of the test substance. The corresponding treatment (25 uL/ear) was applied to the dorsal surface of both ears for three consecutive days. Three days after the last exposure, all animals were injected with 0.25 mL of sterile phosphate buffered solution containing 3H-methyl thymidine and subsequently euthanized The auricular lymph nodes were removed to visually estimate the relative size and any abnormalities. The nodes were pooled to measure the amount of operative DNA by disintegrations per minute (DPM). The stimulation index (SI) was calculated for each group. Consideration was given to the EC3 value. The 6-month reliability check with alpha-hexylcinnamicaldehyde indicated that the LLNA is an appropriate model for evaluating dermal sensitization at the test facility. Observations for mortality (twice daily), body weights (Day 1 and 6), clinical signs (one daily), and irritation (once daily) were performed as well. Mean DPM/animal values for the 25%, 50%, and 100% test article concentrations were 603, 1004, and 370 respectively. The control mean DPM/animal value was 441. The SI values for the 25, 50, and 100% concentrations were 1.4, 2.3, and 0.8 respectively. No EC3 value was calculated because all SI values were <3. There was very slight erythema of the ears in animals treated with the 50% and 100% concentrations of the test material, but was considered not toxicologically significant. No mortality, clinical signs of toxicity, or significant body weight change were observed. All auricular lymph nodes were considered normal in size. Based on the results of the test, extracts of the test article showed no evidence of dermal sensitization.