2,2-dimethylpropane-1,3-diyl dibenzoate

Skin sensitisation

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:

other: Guideline study and GLP

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2014

Report date:

2014

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

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

Deviations:

yes

Remarks:

The modification refer to the measurement of cell proliferation by cell counting instead of radioactive labelling. In addition, the acute inflammatory skin reaction is determined to discriminate specific from non-specific activation of immune system).

Principles of method if other than guideline:

Modified LLNA (IMDS = Integrated Model for the Differentiation of Skin Reactions): The modification refers to the measurement of cell proliferation by cell counting instead of radioactive labelling. In addition, the acute inflammatory skin reaction is determined to discriminate specific from non-specific activation of immune system, as also recommended in the update of OECD TG 429.

GLP compliance:

yes

Type of study:

mouse local lymph node assay (LLNA)

Test material

Test material information

Constituent 1

Reference substance name:

2,2-dimethylpropane-1,3-diyl dibenzoate

EC Number:

224-081-9

EC Name:

2,2-dimethylpropane-1,3-diyl dibenzoate

Cas Number:

4196-89-8

Molecular formula:

C19H20O4

IUPAC Name:

3-(benzoyloxy)-2,2-dimethylpropyl benzoate

impurity 1

Reference substance name:

3-hydroxy-2,2-dimethylpropyl benzoate

Cas Number:

5522-92-9

Molecular formula:

C12H16O3

IUPAC Name:

3-hydroxy-2,2-dimethylpropyl benzoate

Test material form:

solid: crystalline

Details on test material:

Batch No.: GSOH 140121

In vivo test system

Test animals

Species:

mouse

Strain:

NMRI

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Weight at study initiation: 29.0-29.3 g
- Housing: singly during study
- Diet ad libitum
- Water ad libitum
- Acclimation period: 6 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22
- Humidity (%): 40-70
- Air changes (per hr): 10
- Photoperiod (hrs dark / hrs light): 12/12

Study design: in vivo (LLNA)

Vehicle:

other: A/OO = acetone/olive oil (4:1)

Concentration:

0% (vehicle control), 2 %, 10 %, 50 %, and positve control 40 % alpha cinnamic aldehyde

No. of animals per dose:

6

Details on study design:

The test item in the formulation, the positive control in the formulation or the vehicle were applied epicutaneousely onto the dorsal part of both ears of the animals. This treatment was repeated on three consecutive days. The volume administered was 25 µl/ear. The used concentrations were based on the experience with this test system and the properties of the test item.
The animals were anaesthetized by inhalation of carbon dioxide and sacrificed one day after the last application. The appropriate organs were then removed .Lymphatic organs (the auricular lymph nodes) were transferred into physiological saline
Investigations:
- weight of lymph nodes
- cell counts in lymph nodes
- stimulation index is calculated by dividing the absolute number of weight or cell count of the substance-treated lymph nodes by the vehicle treated ones
- ear swelling
- ear weight
- body weight

Positive control substance(s):

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

Statistics:

When it was statistically reasonable, the values from treated groups were compared with those from the control group by one-way Analysis of Vartiance (ANOVA) when the variances are considered homogenous according to a homogeneity testing like Cochran's test. Alternatively, if the variances are considered to be heterogeneous a non-parametric Kruskal-Wallis test has been used at significance levels of 5%. Two sided multiple test procedures were done according to Dunnett or Bonferroni-Holm, respectively. Outlying values in the LN weights were eliminated at a probability level of 99% by Nalimov's method. In addition, for the LLNA/IMDS the smallest significant differences in the means were calculated by Sheffe's method, which can be used for both equal and unequal sample sizes.

Results and discussion

In vivo (LLNA)

Results

Results 1

Key result

Parameter:

SI

Remarks on result:

other: see Remark

Remarks:

It has to be clarified that the positive levels are exclusively defined for the NMRI mice used for this study: Compared to vehicle treated animals, none of the parameter measured in the test substance groups i.e. cell counts, weights of the draining lymph nodes, ear weights and ear swelling reached or exceeded the 'positive level' defined for this assay. The determined cell count indices were 1.0 (vehicle), 1.13, 1.37 and 1.33. Thus, the positive level, which is 1.4 for cell count index was never reached or exceeded. The respective value for the positive controls is 1.68.

Results 2

Key result

Parameter:

other: disintegrations per minute (DPM)

Remarks on result:

other: Modified LLNA. Measurement of cell counts instead of radioactive labelling.

Any other information on results incl. tables

It has to be clarified that the positive levels are exclusively defined for the NMRI mice used for this study:

The positive level of ear swelling which is 2E-02 mm increase, i.e. about 10 % of the control values, has not been reached or exceeded in any dose group of the test item whereas the respective value of the positive control has shown a statistical significant increase.

Body weights of the animals was not affected by any treatment.

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

The LLNA was negative. 2,2-dimethylpropane-1,3-diyl dibenzoate shows no sensitizing potential in the modified Local Lymph Node Assay (IMDS) in female NMRI mice after dermal application of up to and including a 50 % concentration of 2,2-dimethylpropane-1,3-diyl dibenzoate.

Executive summary:

A LLNA/IMDS was carried out in female NMRI mice after epicutaneous application of formulation containing 0%, 2 %, 10 %, or 50 % of 2,2-dimethylpropane-1,3-diyl dibenzoate dissolved in aceton/olive oil (4:1) for 3 consecutive days onto both ears of the animals. The study was conducted according to OECD TG 429 and OECD TG 406 under GLP conditions. The result show no sensitizing potential in the modified Local Lymph Node Assay (IMDS) in female NMRI mice after dermal application of up to and including a 50 % concentration of 2,2-dimethylpropane-1,3-diyl dibenzoate. Additionally, no indication for a non-specific (irritant) activation by the test item was detected. The positive control hexyl cinnamic aldehyde (CAS No. 101-86-0) was functional for each parameter (Vohr 2014).