N,N-bis(trimethylsilyl)aminopropylmethyldiethoxysilane

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

1 May 2003 to 3 June 2003

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2003

Report date:

2003

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

2002

Deviations:

no

GLP compliance:

yes

Type of study:

mouse local lymph node assay (LLNA)

Test material

Test material information

Constituent 1

Reference substance name:

-

EC Number:

445-890-5

EC Name:

-

Cas Number:

201290-01-9

Molecular formula:

C14H37NO2Si3

IUPAC Name:

7-ethoxy-2,2,7-trimethyl-3-(trimethylsilyl)-8-oxa-3-aza-2,7-disiladecane

Test material form:

liquid

Specific details on test material used for the study:

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: at room temperature in the dark
- Stability under test conditions: stable
- Solubility and stability of the test substance in the solvent/vehicle: a vehicle trial showed that the test substance forms a clear liquid at 50% v/v in dimethylformamide (DMF).
- Reactivity of the test substance with the solvent/vehicle of the cell culture medium: The absorption of the test substance was not determined. Chemical analysis of the homogeneity, stability and purity of the test substance was not undertaken.

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing: the test material was dissolved in dimethylformamide

In vivo test system

Test animals

Species:

mouse

Strain:

CBA/Ca

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: Harlan UK Ltd., England
- Females (if applicable) nulliparous and non-pregnant: [not specified]
- Microbiological status of animals, when known: no data
- Age at study initiation: 8 - 12 weeks old
- Weight at study initiation: 16.5 to 22.9 g
- Housing: individually in polycarbonate cages with woodflake bedding
- Diet (e.g. ad libitum): standard laboratory rodent diet, ad libitum
- Water (e.g. ad libitum): drinking water, ad libitum
- Acclimation period: 6 days
- Indication of any skin lesions: none reported

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21 +/- 2°C
- Humidity (%): 40 - 70%
- Air changes (per hr): no data
- Photoperiod (hrs dark / hrs light): 12/12

Study design: in vivo (LLNA)

Vehicle:

dimethylformamide

Concentration:

Preliminary study: 25 and 50% v/v, and 100% in DMF
Main study: 25 and 50% v/v, and 100% in DMF

No. of animals per dose:

4 mice per dose

Details on study design:

PRE-SCREEN TESTS:
One female was treated at one of three concentrations of the test substance. The mice were treated by daily application of 25 µl of appropriate concentration of the test substance to the dorsal surface of each ear for three consecutive days. The test substance was applied to the dorsal surface of each ear using an automatic micropipette. The test substance was spread over the entire dorsal surface of the ear using the tip of the pipette.

MAIN STUDY
Groups of four mice were treated at one of three concentrations of the test substance. The mice were treated by daily application of 25 µl of the appropriate concentration of the test substance to the dorsal surface of each ear for three consecutive days. The test substance was applied to the dorsal surface of each ear using an automatic micropipette. The test substance was spread over the entire dorsal surface of the ear using the tip of the pipette. A further group of four mice received the vehicle alone in the same manner. Five days following the first topical application of the test substance (day 6) all mice were injected via the tail vein with 250µl of phosphate buffered saline containing ³H-methyl thymidine giving a normal 20 µCi to each mouse. The injection into the tail vein was carried out using a plastic syringe and needle after the mouse had been heated in a warming chamber.

ANIMAL ASSIGNMENT AND TREATMENT
- Criteria used to consider a positive response: The test substance is considered to be a sensitiser if at least one concentration of the test substance results in a three-fold greater increase in ³H-methyl thymidine incorporation compared to control values (SI > 3).

OBSERVATIONS
Daily observations for clinical signs of toxicity. The ears were also examined for signs of irritation. Body weights of mice in the preliminary and main study were recorded on arrival, day 1 and prior to termination.

TERMINAL STUDIES
- Termination: 5 hours after administration of ³H-methyl thymidine on day 6 all mice were killed and the draining auricular lymph nodes excited and pooled for each experimental group. 1.0. ml of phosphate buffered saline was added to the pooled lymph nodes. The animals were then discarded and no further investigations were carried out.
- Preparation of single cell suspensions: A single cell suspension of lymph node cells (LNC) was prepared by gentle mechanical disaggregation through a stainless steel gauze. The pooled LNC were then washed, pelleted for 10 minutes and resuspended. The cells were washed twice again and resuspended in 3 ml trichloroacetic acid (TCA) following the final wash.
- Determination of incorporated ³H-methyl Thymidine: After an overnight incubation with TCA the precipitate was recovered by centrifugation and resuspended. ³H-methyl Thymidine incorporation was determined by β-scintillation counting. The proliferative response of LNC was expressed as radioactive disintegrations per minute per lymph node and as the ration of ³H-methyl Thymidine incorporation into LNC of test nodes relative to that recorded for control nodes.

Positive control substance(s):

not specified

Results and discussion

In vivo (LLNA)

Results

Results 1

Key result

Parameter:

SI

Value:

11

Test group / Remarks:

25% v/v in DMF

Results 2

Key result

Parameter:

SI

Value:

17.5

Test group / Remarks:

50% v/v in DMF

Results 3

Key result

Parameter:

SI

Value:

27.1

Test group / Remarks:

100%

Results 4

Key result

Parameter:

EC3

Value:

10.96

Remarks on result:

other: EC3 value was estimated based on SI data.

Cellular proliferation data / Observations:

CELLULAR PROLIFERATION DATA:
The test/control ratios obtained for 25, 50 and 100 % were 11.0, 17.5 and 27.1, respectively. As a test/ control ratio of 3 or more was recorded for all of the concentrations tested , the test substance was considered to have the potential to cause skin sensitisation (delayed contact hypersensitivity).

DETAILS ON STIMULATION INDEX CALCULATION: ³H-methyl Thymidine incorporation was determined by β-scintillation counting. The proliferative response of LNC was expressed as radioactive disintegrations per minute per lymph node and as the ration of ³H-methyl Thymidine incorporation into LNC of test nodes relative to that recorded for control nodes.

CLINICAL OBSERVATIONS:
- Preliminary study: No deaths occurred during the study. Post-dose wet fur around the cranial region of all the animals was observed throughout the three groups from day 1. The animals appeared normal by day 4 post-dosing. On the basis of the results, 100% was considered a suitable high dose level for the main study.
- Main study: No deaths occurred during the study period. No clinical signs of toxicity were noted in any of the test animals. Post-dose wet fur around the cranial region of all the animals was observed throughout the three groups from day 1. The animals appeared normal by day 4 post-dosing.

BODY WEIGHTS
- Preliminary study: Bodyweight loss was recorded in one animal in group 2
- Main study: The expected body weight gain was observed in all the animals.

Applicant's summary and conclusion

Interpretation of results:

Category 1B (indication of skin sensitising potential) based on GHS criteria

Conclusions:

In the skin sensitisation study, conducted according to OECD TG 429 and in compliance with GLP, the test material, N,N-bis(trimethylsilyl)aminopropylmethyldiethoxysilane, was reported to be a skin sensitiser.