Dialuminium zinc tetraoxide

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

19 - 26 Oct 2011

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP - Guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2011

Report date:

2011

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

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

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.42 (Skin Sensitisation: Local Lymph Node Assay)

Test guideline 3

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.2600 (Skin Sensitisation)

GLP compliance:

yes (incl. QA statement)

Remarks:

GYEMSZI, National Institute for Quality and Organizational Development in Healthcare and Medicines, National Institute for Pharmacy

Type of study:

mouse local lymph node assay (LLNA)

Test material

Test material information

Constituent 1

Reference substance name:

Dialuminium zinc tetraoxide

EC Number:

235-101-0

EC Name:

Dialuminium zinc tetraoxide

Cas Number:

12068-53-0

Molecular formula:

ZnAl2xO(3x+1), x = 1 – 9

IUPAC Name:

dialuminum; zinc(2+); oxygen(2-)

Details on test material:

- Name of test material (as cited in study report): D-PrisM
- Physical state: beige (off-white) granulate
- Analytical purity: 100%
- Composition of test material, percentage of components: dialuminium zinc tetraoxide (zinc aluminate)
- Lot/batch No.: PC080519/1
- Expiration date of the lot/batch: 01.06.2018
- Storage condition of test material: room temperature

In vivo test system

Test animals

Species:

mouse

Strain:

other: CBA/Ca (SPF)

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: TOXI-COOP ZRT., Budapest, Cserkesz, Hungary
- Age at study initiation: 11 - 13 weeks (dose range finding test), 9 - 11 weeks (main study)
- Weight at study initiation: 16.7 - 21.3 g (main study)
- Housing: 4 animals per cage in Type II propylene / polycarbonate cages on deep wood sawdust bedding (Lignocel® Hygienic Animal Bedding, J. Rettenmaier & Söhne GmbH + Co. KG, Germany)
- Diet: ssniff® SM R/M-Z+H complete diet, ssniff Spezialdiäten GmbH, Soest, Germany, ad libitum
- Water: tap water, ad libitum
- Acclimation period: 7 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3
- Humidity (%): 30 - 70
- Photoperiod (hrs dark / hrs light): 12 / 12

IN-LIFE DATES: From: 12 Oct 2011 To: 24 Oct 2011

Study design: in vivo (LLNA)

Vehicle:

other: N,N-Dimethylformamide (DMF)

Concentration:

10, 25 and 50% (w/v) (suspension)

No. of animals per dose:

4

Details on study design:

RANGE FINDING TESTS:
- Compound solubility: The test item was insoluble in all vehicles tested (acetone:olive oil (4:1 mixture), N,N-dimethylformamide (DMF), methyl ethyl ketone, dimethyl sulfoxide). Thus, the test substance suspension was prepared in DMF as DMF is one of the most preferred vehicles generally used for insoluble materials (such as medical devices) and it does not evaporate from the skin surface as rapidly as an acetone:olive oil mixture (4:1). The maximum available concentration was 50% (w/v). Although the suspension prepared in this way sedimented rapidly applicability on the ears of animals (with continuous stirring until application) it was found to be appropriate.
- Irritation:
Groups of 2 mice were treated with 50 or 25% test item solution. All animals were observed for any clinical signs of systemic toxicity or local irritation at the application site. Body weights were recorded prior to the first treatment (Day 1) and prior to termination (Day 6). Both ears of each mouse were observed for erythema and scored. Measurement of ear thickness was taken using digital micrometer on Day 1 (pre-dose), Day 3 (approximately 48 hours after the first dose) and Day 6.
No mortality or any signs of systemic toxicity were observed. No significant signs of irritation were observed as indicated by an erythema score ≥ 3 and/or an increased ear thickness of ≥ 25% on any day of measurement (50%: 0.23 mm on day 1 and 3, 0.24 on day 6; 25%: 0.23 mm on day 1, 3 and 6). According to this, a concentration of 50 % (w/v) was selected
as the highest test concentration in the main test.
- Lymph node proliferation response: Lymph node proliferation was not assessed in the range-finding test.

MAIN STUDY
ANIMAL ASSIGNMENT AND TREATMENT
- Name of test method: 3H-methyl thymidine incorporation determined by ß-scintillation
- Criteria used to consider a positive response:
The test item was considered as a skin sensitizer, if the following criteria were fulfilled:
- exposure to at least one concentration of the test item resulted in an at least 3-fold or greater stimualtion index in test animals compared to control mice.
- the data are compatible with a conventional dose response, although allowance must be made (especially at high topical concentrations) for either local toxicity or immunological suppression.
The EC3 value (dose calculated to induce a stimulation index of 3) of the test item was not calculated.

TREATMENT PREPARATION AND ADMINISTRATION:
25 µL of the test item was applied on the dorsal surface of each ear of each dose. The application was repeated on Day 2 and 3. Local irritation reactions were assessed. On day 6, an injection of 250 µL phosphate buffered saline containing 20 µCi of 3H-methyl thymidine was made into the tail vein of each experimental mouse. 5 ± 0.5 h later, animals were sacrificed and the draining auricular lymph node of each ear was excised. A single cell suspension of lymph node cells, pooled according to groups, was prepared and collected in disposable tubes by gentle mechanical disaggregating of the lymph nodes through a cell strainer using the plunger of a disposable syringe. After repeated washings, cells were precipitated with 5% trichloracetic acid at 2 - 8 °C for approx. 18 hrs.

CLINICAL OBSERVATIONS:
All animals were observed at least once a day throughout the study period for any clinical signs, including systemic toxicity and local irritation. Irritation was monitored by erythema scoring (during the whole test) according to the scoring scale defined in OECD GL 429 and recorded for each animal individually .

Positive control substance(s):

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

Results and discussion

Positive control results:

The positive control substance induced an enlargement of the lymph nodes with a significant lymphoproliferative response as shown by a stimulation index value of 15.0.

In vivo (LLNA)

Results

Results 1

Parameter:

SI

Value:

0.7

Test group / Remarks:

10%

Results 2

Parameter:

SI

Value:

1

Test group / Remarks:

25%

Results 3

Parameter:

SI

Value:

0.7

Test group / Remarks:

50%

Results 4

Parameter:

other: disintegrations per minute (DPM)

Remarks on result:

other: No significant or dose-dependent increase in the DPM values was observed in the test groups compared to the controls. The determined DPM values per group were 4761, 3106, 4761 and 3491 for the control, 10%, 25% and 50% groups, respectively.

Any other information on results incl. tables

No significant effects on the body weight were observed in any treatment group. In addition, no mortality or symptoms of systemic toxicity were determined.

Applicant's summary and conclusion

Interpretation of results:

other: CLP/EU GHS criteria are not met, no classification required according to Regulations (EC) No 1272/2008

Conclusions:

CLP: not classified