N-{3-[dimethoxy(methyl)silyl]propyl}butan-1-amine

Skin irritation / corrosion

Administrative data

Endpoint:

skin corrosion: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

16 - 18 Jan 2018

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2018

Report date:

2019

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 431 (In Vitro Skin Corrosion: Reconstructed Human Epidermis (RHE) Test Method)

Version / remarks:

29 July, 2016

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

Bayerisches Landesamt für Gesundheit und Lebensmittelsicherheit, Schwabach, Germany

Test material

Test material information

Constituent 1

Reference substance name:

N-{3-[Dimethoxy(methyl)silyl]propyl}butan-1-amine

EC Number:

601-747-0

Cas Number:

120939-52-8

Molecular formula:

C10H25NO2Si

IUPAC Name:

N-{3-[Dimethoxy(methyl)silyl]propyl}butan-1-amine

In vitro test system

Test system:

human skin model

Source species:

human

Cell type:

non-transformed keratinocytes

Cell source:

other: EpiDerm™

Details on test system:

RECONSTRUCTED HUMAN EPIDERMIS (RHE) TISSUE
- Model used: reconstituted three-dimensional human skin model EPIDERM™
- Tissue batch number(s): 25874 and 25848

TEMPERATURE USED FOR TEST SYSTEM
- Temperature used during treatment / exposure: 37 ± 1 °C
- Temperature of post-treatment incubation: 37 ± 1 °C

REMOVAL OF TEST MATERIAL AND CONTROLS
- Volume and number of washing steps: Tissue was gently rinsed about 20 times with PBS to remove any residual test item. Excess PBS was removed by gently shaking the insert and blotting bottom with blotting paper

MTT DYE USED TO MEASURE TISSUE VIABILITY AFTER TREATMENT / EXPOSURE
- MTT concentration: MTT stock solution (i.e., 5 mg/mL MTT in PBS) diluted 1 + 4 with Dulbecco’s Modified Eagle Medium (DMEM)-based medium (final concentration 1 mg/mL)
- Incubation time: 3 h
- Spectrophotometer: Yes, plate spectrophotometer
- Wavelength: 570 nm
- Filter: Not reported

CONTROL TISSUES USED IN CASE OF MTT DIRECT INTERFERENCE
- Killed tissues: The mixture of 50 µL test item per 1 mL MTT medium showed reduction of MTT as compared to the solvent. The mixture turned blue/purple. For quantitative correction of results, the part of absorption due to the non-specific reduction of MTT (NSMTT) was determined by using killed tissues. Therefore, two tissues per treatment period were treated with the test item (KT) or left untreated (KU), respectively. NSMTT was calculated relative to the negative control of living tissues (NK) per treatment period according to the following formula:
NSMTT = [(ODKT – ODKU / ODNK] * 100

NSMTT was ≤ 30% relative to the negative control of living epidermis. In the 3 min experiment NSMTT was -18.42%, in the 60 min experiment 10.90%. This means that the test item was washed away almost completely before the addition of the MTT solution. The true MTT metabolic conversion (TODTT) of the test item treated living tissues (TM) was corrected for each treatment period according to the following formula:
TODTT = ODTM - (ODKT - ODKU)

The mixture of 50 µL test item per 300 µL Aqua dest. and per 90 µL isopropanol showed no colouring as compared to the solvent. Therefore, no additional controls for non-specific colouring potential were included.

FUNCTIONAL MODEL CONDITIONS WITH REFERENCE TO HISTORICAL DATA
- Viability: 1.71 ± 0.9 for lot 25874; 1.257 ± 0.111 for lot 25848
- Barrier function: 5.28 h for lot 25874; 7.34 h for lot 25848
- Contamination: Screened for HIV-1, hepatits B and C, bacteria, yeast, and other fungi. No contaminates were detected.

NUMBER OF REPLICATE TISSUES: 2

PREDICTION MODEL / DECISION CRITERIA
A substance was considered corrosive if tissue viabilities were: a) < 50% after 3 min exposure; b) ≥ 50% after 3 min exposure AND < 15% after 60 min exposure; c). < 25% after 3 min exposure ; OR ≥ 25% after 3 min exposure. A substance was considered noncorrosive if tissue viabilities were ≥ 50% after 60 min exposure AND ≥ 15% after 60 min exposure.

The test meets acceptance criteria if:
- mean absolute OD570 nm of the two negative control tissues of every treatment period is between 0.8 and 2.8,
- mean relative tissue viability of the two positive control tissues of the 60 min treatment period is < 15%,
- coefficient of variation (CV) (in the range of 20 – 100% viability) between two tissues treated identically is ≤ 30%.

Control samples:

yes, concurrent negative control

yes, concurrent positive control

Amount/concentration applied:

TEST MATERIAL
- Amount applied: 50 µL

NEGATIVE CONTROL
- Amount applied: 50 µL

POSITIVE CONTROL
- Amount applied: 50 µL

Duration of treatment / exposure:

3 min and 60 min

Number of replicates:

The test was performed on a total of 4 tissues per dose group, 2 replicates for each treatment period (3 min and 60 min exposure time).

Results and discussion

In vitro

Results

Results 1

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

3 min treatment

Value:

104.1

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Results 2

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

60 min treatment

Value:

22.2

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Other effects / acceptance of results:

- OTHER EFFECTS:
- Direct-MTT reduction: The test item showed non-specific MTT-reducing potential.
- Colour interference with MTT: The test item showed no colour interference.

DEMONSTRATION OF TECHNICAL PROFICIENCY: The controls confirmed the validity of the study. The mean OD570nm of the two negative control tissues was ≥ 0.8 and ≤ 2.8 for each exposure period. The mean relative tissue viability (% negative control) of the positive control was < 15% (1.5%) after 60 min treatment. The coefficient of variation (CV) (in the range of 20 – 100% viability) of replicate tissues of all dose groups was ≥ 30% (20.1% - 21.8%). Tabulated test acceptance criteria and historical control data are presented in Attachment 1.

ACCEPTANCE OF RESULTS:
- Acceptance criteria met for negative control: yes
- Acceptance criteria met for positive control: yes
- Acceptance criteria met for variability between replicate measurements: yes

Applicant's summary and conclusion

Interpretation of results:

other: non-corrosive according to Regulation (EC) No. 1272/2008

Conclusions:

Under the conditions of the conducted test, the test substance did not possess corrosive properties towards reconstructed human epidermis tissue in the EpiDerm™ model.