Allyltrimethylsilane

Endpoint summary

Description of key information

In vitro skin irritation (OECD 431), human skin model test (EpiDerm): not corrosive

In vitro skin irritation (OECD 439), human skin model test (EpiSkin): not irritating

Eye irritation (OECD 437), bovine cornea in vitro: not irritating

Key value for chemical safety assessment

Skin irritation / corrosion

Link to relevant study records

Reference

Reference 1

Endpoint:

skin corrosion: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

06 to 08 March 2018

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

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)

Test system:

human skin model

Source species:

human

Cell type:

non-transformed keratinocytes

Cell source:

other: Human donors

Source strain:

not specified

Vehicle:

unchanged (no vehicle)

Details on test system:

RECONSTRUCTED HUMAN EPIDERMIS (RHE) TISSUE
- Model used: EpiDerm™ Reconstructed Human Epidermis Model Kit
- Tissue batch number(s): 28686
- Delivery date: 05 March 2019
- Date of initiation of testing: 06 March 2019

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

REMOVAL OF TEST MATERIAL AND CONTROLS
-Volume and number of washing steps: Rinsing was achieved by filling and emptying each tissue under a constant soft stream of Dulbecco's Phosphate Buffered Saline (DPBS) (without Ca++ Mg++) for approximately 40 seconds, to gently remove any residual test item. Excess DPBS was removed by blotting the bottom of the tissue insert with tissue paper.

MTT DYE USED TO MEASURE TISSUE VIABILITY AFTER TREATMENT / EXPOSURE
- MTT concentration: 1.0 mg/mL MTT solution
- Incubation time: 3 hours
- Spectrophotometer: yes
- Wavelength: 570 nm
- Filter: Yes
- Filter bandwidth: 10 nm

NUMBER OF REPLICATE TISSUES: The test was performed on a total of 6 tissues for each test item and for the negative and positive controls, with 2 replicates for each treatment period (3 min and 60 min).

NUMBER OF INDEPENDENT TEST SEQUENCES / EXPERIMENTS TO DERIVE FINAL PREDICTION:
Three independent experiments were conducted with the following exposure periods: 3 min and 60 min

PREDICTION MODEL / DECISION CRITERIA
A substance was considered corrosive if tissue viabilities were < 50% after 3 min exposure or if tissue viabilities were ≥ 50% after 3 min exposure AND < 15% after 60 min exposure. If a substance was considered corrosive, it was classified as Category 1A if tissue viabilities were < 20% after 3 min exposure. It was classified as Category 1B/1C if tissue viabilities were ≥ 25% after 3 min exposure. A substance was considered non-corrosive if tissue viabilities were ≥ 50% after 3 min exposure AND ≥ 15% after 60 min exposure.

The test meets acceptance criteria if:
- The mean OD570 of the two negative control tissues was between ≥ 0.8 and ≤ 2.8 for each exposure time.
- The mean relative tissue viability of the 60-minute positive control is < 15%.

In the range 20 and 100% viability, the Coefficient of Variation between tissue replicates should be ≤ 30%.

Control samples:

yes, concurrent negative control

yes, concurrent positive control

Amount/concentration applied:

TEST MATERIAL
- Amount(s) applied: 50 µL
- Concentration: undiluted

NEGATIVE CONTROL
- Amount(s) applied: 50 µL
- Concentration: undiluted

POSITIVE CONTROL
- Amount(s) applied: 50 µL
- Concentration: 8.0N Potassium Hydroxide

Duration of treatment / exposure:

3 min or 60 min

Duration of post-treatment incubation (if applicable):

3 hours

Number of replicates:

The test was performed on a total of 6 tissues for each test item, negative control, and positive control, 2 replicates for each treatment period (3 min and 60 min).

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

3 min

Value:

87.5

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

60 min

Value:

98.4

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Other effects / acceptance of results:

- OTHER EFFECTS:
- Visible damage on test system: The test item showed no visible damage to the test system.
- Direct-MTT reduction: The test item showed no non-specific MMT-reducing potential.
- Colour interference with MTT: The solution containing the test item did not become colored. This was taken to indicate the test item did not have the potential to cause color interference.

DEMONSTRATION OF TECHNICAL PROFICIENCY: The test item showed no corrosive effects. All test acceptance criteria were met. Positive and negative controls responded appropriately. The maximum inter-tissue viability difference of replicate tissues fell within the acceptance criteria.

ACCEPTANCE OF RESULTS:
- Acceptance criteria met for negative control: The mean OD570 for the negative control treated tissues was 2.469 for the 3-Minute exposure period and 2.314 for the 60-Minute exposure period. The negative control acceptance criteria were therefore satisfied.
- Acceptance criteria met for positive control: The relative mean tissue viability for the positive control treated tissues was 2.5% relative to the negative control following the 60-Minute exposure period. The positive control acceptance criterion was therefore satisfied.
- Acceptance criteria met for variability between replicate measurements: In the range 20 to 100% viability the Coefficient of Variation between the two tissue replicates of each treatment group did not exceed 30%. The acceptance criterion was therefore satisfied.

Interpretation of results:

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

Conclusions:

In an in vitro skin corrosion human skin model test according to OECD guideline 431 and in compliance with GLP, cell viabilities of 98.4% after 60 minutes of exposure, and 87.5% after 3 minutes of exposure were measured when compared to the untreated control. Therefore, the test item should not be classified as corrosive.

Reference 2

Endpoint:

skin irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

10 to 15 April 2019

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 439 (In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method)

Version / remarks:

28 July 2015

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.46 (In Vitro Skin Irritation: Reconstructed Human Epidermis Model Test)

Version / remarks:

06 July 2012

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

Department of Health of the Government of the United Kingdom, Medicines and Healthcare Products, Regulatory Agency, United Kingdom

Test system:

human skin model

Source species:

human

Cell type:

non-transformed keratinocytes

Cell source:

other: Human donors

Source strain:

not specified

Vehicle:

unchanged (no vehicle)

Details on test system:

RECONSTRUCTED HUMAN EPIDERMIS (RHE) TISSUE
- Model used: EpiSkin Laboratories, Lyon, France
- Tissue batch number(s): 19-EKIN-015
- Delivery date: 09 April 2019
- Date of initiation of testing: 10 April 2019

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

REMOVAL OF TEST MATERIAL AND CONTROLS
-Volume and number of washing steps: At the end of the exposure period, each tissue was removed from the well using forceps and rinsed using a wash bottle containing DPBS with Ca++ and Mg++. Rinsing was achieved by filling and emptying each tissue insert for approximately 40 seconds using a constant soft stream of DPBS to gently remove any residual test item.

MTT DYE USED TO MEASURE TISSUE VIABILITY AFTER TREATMENT / EXPOSURE
- MTT concentration: A 3 mg/mL MTT stock solution was prepared in DPBS. The stock solution was diluted to 0.3 mg/mL with assay medium when required.
- Incubation time: 3 hours
- Spectrophotometer: yes
- Wavelength: 570 nm
- Filter: yes
- Filter bandwidth: 10 nm

NUMBER OF REPLICATE TISSUES: triplicate

PREDICTION MODEL / DECISION CRITERIA (choose relevant statement)
The irritant potential was predicted from the relative mean tissue viabilities compared to the negative control. If the mean relative tissue viability is > 50% the test material is not considered an irritant. If the mean relative tissue viability is ≤ 50%, the test material is considered an irritant.

The test meets acceptance criteria if:
- mean absolute OD570 of the negative control tissues is ≥ 0.6 and ≤ 1.5 and the SD value of the percentage viability is ≤ 18%
- mean relative tissue viability of the positive control tissues is ≤ 40% relative to the negative control treated tissues, and the standard deviation (SD) value of the percentage viability is ≤ 18%
- the standard deviation calculated from individual percentage tissue viabilities of the three identically treated tissues is ≤ 18%

Control samples:

yes, concurrent negative control

yes, concurrent positive control

Amount/concentration applied:

TEST MATERIAL
- Amount(s) applied: 10 µL
- Concentration: undiluted

NEGATIVE CONTROL
- Amount(s) applied: 10 µL
- Concentration: undiluted

POSITIVE CONTROL
- Amount(s) applied: 10 µL
- Concentration: 5% Sodium dodecyl sulphate

Duration of treatment / exposure:

15 minutes

Duration of post-treatment incubation (if applicable):

42 hr

Number of replicates:

3

Irritation / corrosion parameter:

% tissue viability

Value:

85

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Other effects / acceptance of results:

- OTHER EFFECTS:
- Visible damage on test system: The test item showed no visible damage to the test system.
- Direct-MTT reduction: The MTT solution containing the test item did not turn blue or purple which indicated that the test item did not directly reduce MTT.
- Colour interference with MTT: The solution containing the test item was colorless. It was therefore unnecessary to run color correction tissues.

DEMONSTRATION OF TECHNICAL PROFICIENCY: The test item showed no irritative effects. Positive and negative controls responded appropriately. The maximum inter-tissue viability difference of replicate tissues fell within the acceptance criteria. Positive and negative controls responded appropriately. The maximum inter-tissue viability difference of replicate tissues fell within the acceptance criteria.

ACCEPTANCE OF RESULTS:
- Acceptance criteria met for negative control: The mean OD570 for the negative control treated tissues was 0.868 and the standard deviation value of the viability was 7.8%. The negative control acceptance criteria were therefore satisfied.
- Acceptance criteria met for positive control: The relative mean tissue viability for the positive control treated tissues was 17.5% relative to the negative control treated tissues and the standard deviation value of the viability was 9.5%. The positive control acceptance criteria were therefore satisfied.
- Acceptance criteria met for variability between replicate measurements: The standard deviation calculated from individual tissue viabilities of the three identically test item treated tissues was 0.7%. The test item acceptance criterion was therefore satisfied.

Interpretation of results:

other: CLP/EU GHS criteria are not met, the test substance is considered to be not corrosive accordi ng to Regulations (EC) No 1272/2008.

Conclusions:

In an in vitro skin irritation human skin model test (EpiDerm) according to OECD guideline 439 and in compliance with GLP, a cell viability of 85.0% after a 15-Minute exposure period and 42-Hour post-exposure incubation period was measured. Therefore, the test item does not meet the criteria for classification.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Eye irritation

Link to relevant study records

Reference

Endpoint:

eye irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

16 April 2019

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 437 (Bovine Corneal Opacity and Permeability Test Method for Identifying i) Chemicals Inducing Serious Eye Damage and ii) Chemicals Not Requiring Classification for Eye Irritation or Serious Eye Damage)

Version / remarks:

09 October 2017

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

Department of Health of the Government of the United Kingdom, Medicines and Healthcare Products, Regulatory Agency, United Kingdom

Species:

cattle

Strain:

not specified

Details on test animals or tissues and environmental conditions:

SOURCE OF COLLECTED EYES
- Source: obtained from a local abattoir as a by-product from freshly slaughtered animals
- Number of animals: not reported
- Characteristics of donor animals: adult cattle (typically 12 to 60 months old)
- Storage, temperature and transport conditions of ocular tissue: The eyes were excised by an abattoir employee after slaughter, and were placed in Hanks’ Balanced Salt Solution (HBSS) supplemented with antibiotics (penicillin at 100 IU/mL and streptomycin at 100 µg/mL). They were transported to the test facility over ice packs on the same day of slaughter.
- Time interval prior to initiating testing: The corneas were prepared immediately on arrival.
- Indication of any existing defects or lesions in ocular tissue samples: All eyes were macroscopically examined before and after dissection. Only corneas free of damage were used.

Vehicle:

unchanged (no vehicle)

Controls:

yes, concurrent positive control

yes, concurrent negative control

Amount / concentration applied:

TEST MATERIAL
- Amount(s) applied: 0.75 mL

Duration of treatment / exposure:

10 minutes

Duration of post- treatment incubation (in vitro):

120 minutes

Number of animals or in vitro replicates:

3 corneas for the test item
3 corneas as negative controls treated with 0.9% NaCl
3 corneas as positive controls treated with ethanol

Details on study design:

SELECTION AND PREPARATION OF CORNEAS
The eyes were carefully examined for defects and any defective eyes were discarded.

The tissue surrounding the eyeball was carefully pulled away and the cornea was excised leaving a 2 to 3 mm rim of sclera. The iris and lens were peeled away from the cornea. The isolated corneas were immersed in a dish containing HBSS until they were mounted in Bovine Corneal Opacity and Permeability (BCOP) holders. The anterior and posterior chambers of each BCOP holder were filled with complete Eagle's Minimum Essential Medium (EMEM) without phenol red and plugged. The holders were incubated for 60 minutes. At the end of the incubation period each cornea was examined for defects. Only corneas free of damage were used.

NUMBER OF REPLICATES
3

NEGATIVE CONTROL USED
yes; 0.75 mL sodium chloride 0.9% w/v

POSITIVE CONTROL USED
yes; 0.75 mL ethanol

APPLICATION DOSE AND EXPOSURE TIME
0.75 mL test material for 10 minutes

POST-INCUBATION PERIOD: no

REMOVAL OF TEST SUBSTANCE
- Number of washing steps after exposure period: test substance or the control substance was removed and the epithelium washed at least three times with fresh complete EMEM containing phenol red before a final rinse with complete EMEM without phenol red.
- POST-EXPOSURE INCUBATION: 120 minute incubation at 32 ± 1 °C

METHODS FOR MEASURED ENDPOINTS:
- Corneal opacity: The change in opacity for each cornea was calculated by subtracting the initial opacity reading from the final opacity reading. These values were corrected by subtracting from each the average change in opacity observed for the negative-control corneas. The mean opacity value for each treatment was calculated by averaging the corrected opacity values of each cornea for a given treatment.
- Corneal permeability: The corrected OD492 was calculated by subtracting the mean OD492 of the negative control corneas from the OD492 value of each treated cornea. The OD492 value of each treatment group was calculated by averaging the corrected OD492 values of the treated corneas for the treatment group.

SCORING SYSTEM: The following formula was used to determine the in vitro irritation score (IVIS): IVIS = mean opacity value + (15 x mean permeability OD492 value)

DECISION CRITERIA: The BCOP assay was considered to be valid if the in vitro irritation score obtained with the positive control falls within the two standard deviations of the current historical mean collated during the previous 12 months for this testing facility. For the negative control, BCOP assay was considered to be valid if the in vitro irritation score obtained was less than or equal to the upper limit for background opacity and permeability values calculated from the previous 12 months data for this testing facility.

The following criteria was used for the BCOP Assay: ≤ 3 = not inducing eye damage; > 3; ≤ 55: no prediction can be made; and > 55: inducing serious eye damage. When no prediction for inducing serious eye damage or not inducing eye damage can be made further testing with another suitable method is required.

Irritation parameter:

in vitro irritation score

Value:

0.9

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Other effects / acceptance of results:

OTHER EFFECTS:
- Visible damage on test system: The corneas treated with the test item were clear post treatment and post incubation. The corneas treated with the negative control item were clear post treatment and post incubation. The corneas treated with the positive control item were cloudy post treatment and post incubation.

DEMONSTRATION OF TECHNICAL PROFICIENCY: yes

ACCEPTANCE OF RESULTS:
- Acceptance criteria met for negative control: yes; The negative control gave opacity and permeability values below the established upper limits. The negative control acceptance criteria were therefore satisfied.
- Acceptance criteria met for positive control: yes; The positive control in vitro irritancy score was within the acceptance range. The positive control acceptance criterion was therefore satisfied.

Interpretation of results:

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

Conclusions:

Under the conditions of the test, the test substance was shown to have no irritation or corrosive potential in the bovine corneal opacity and permeability (BCOP) test prediction model. The mean in vitro irritation score was 0.9.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Respiratory irritation

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Skin Irritation

Two in vitro skin irritation/corrosion studies were conducted for allyltrimethylsilane.The OECD 431 study (Lacey, 2019) demonstrated that the test material was not corrosive with cell viabilities measuring 98.4% after 60 minutes of exposure, and 87.5% after 3 minutes. In the OECD 439 study (Lacey, 2019), a cell viability of 85.0% was measured after a 15-minute exposure period and 42-Hour post-exposure incubation period. Therefore, the test item does not meet the criteria for classification.

 

Eye Irritation

In an in vitro eye irritation test (Brown, 2019), the mean in vitro irritation score was 0.9. The study was conducted in accordance with OECD guidelines and was GLP compliant. Therefore, under the conditions of the test, the test substance was shown to have no irritation or corrosive potential.

Justification for classification or non-classification

The available data on skin and eye irritation indicate that allyltrimethylsilane do not meet the criteria for classification according to Regulation (EC) No 1272/2008, and are therefore conclusive but not sufficient for classification.