Benzyl propionate

Endpoint summary

Description of key information

Skin irritation: The test item was not considered to be irritant to skin in the in vitro Human Skin Model Test.
Eye irritation: The test substance was not irritating to rabbit's eye.

Key value for chemical safety assessment

Skin irritation / corrosion

Link to relevant study records

Reference

Endpoint:

skin irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2015-12-11 and 2016-03-22

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:

2015

Deviations:

no

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

2009

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test system:

human skin model

Source species:

other: reconstituted human epidermis

Cell type:

non-transformed keratinocytes

Justification for test system used:

The potential of chemical induced skin irritation is usually determined in vivo in the Draize rabbit skin irritation test as described in OECD guideline 404. Because systemic reactions play a minor role in modulating local skin toxicity potential of chemicals, skin irritation potential may be predicted by in vitro systems, provided they are sufficiently complex to mimic human skin barrier and cell reactivity. In an international prevalidation study performed by ECVAM, the in vitro skin irritation test using the human skin model EpiDerm™ and EpiSkin™ and measurement of cell viability by dehydrogenase conversion of MTT into a blue formazan salt have turned out as a sufficiently promising predictor for skin irritancy potential.

Vehicle:

unchanged (no vehicle)

Details on test system:

RECONSTRUCTED HUMAN EPIDERMIS (RHE) TISSUE
- Model used: Epi-200 SIT kit
- Tissue batch number(s): 23308
- Delivery date: 19 February 2016
- Date of initiation of testing: On day of receipt the pre-incubation phase of the EpiDerm™ tissues started.

TEMPERATURE USED FOR TEST SYSTEM
- Temperature used during treatment / exposure: 35 min at 37°C and 25 min at room temperature
- Temperature of post-treatment incubation: 37°C

REMOVAL OF TEST MATERIAL AND CONTROLS
- Volume and number of washing steps: rinsed with DPBS at least 15 times; submerged in DPBS at least three times; afterwards once again rinsed with sterile DPBS from the inside and outside
- Observable damage in the tissue due to washing: no data
- Modifications to validated SOP: none

MTT DYE USED TO MEASURE TISSUE VIABILITY AFTER TREATMENT / EXPOSURE
- MTT concentration: 1 mg/mL
- Incubation time: 3 hours
- Spectrophotometer: Versamax® Molecular Devices, Softmax Pro, version 4.7.1
- Wavelength: 570 nm

FUNCTIONAL MODEL CONDITIONS WITH REFERENCE TO HISTORICAL DATA
- Viability: OD = 1.673 +/- 0.048
- Barrier function: 8.20 hours
- Contamination: sterile

NUMBER OF REPLICATE TISSUES: 3

NUMBER OF INDEPENDENT TEST SEQUENCES / EXPERIMENTS TO DERIVE FINAL PREDICTION: 1

PREDICTION MODEL / DECISION CRITERIA
- The test substance is considered to be irritant to skin category 2 if the viability is less than or equal to 50%.
- The test substance is considered to be non-irritant to skin if the viability is greater than 50%.

Control samples:

yes, concurrent negative control

yes, concurrent positive control

Amount/concentration applied:

TEST MATERIAL
- Amount applied: 30 µL (47 µL/cm2)

NEGATIVE CONTROL
- Amount applied: 30 µL

POSITIVE CONTROL
- Amount applied: 30 µL
- Concentration: 5%

Duration of treatment / exposure:

60 minutes

Duration of post-treatment incubation (if applicable):

41 hours and 37 minutes

Number of replicates:

3 triplicate tissues (3 wells per tissue)

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

Tissue 1

Value:

106.5

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

Tissue 2

Value:

91.6

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

Tissue 3

Value:

98.9

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Other effects / acceptance of results:

- OTHER EFFECTS:
- Direct-MTT reduction: no
- Colour interference with MTT: no

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

Interpretation of results:

GHS criteria not met

Conclusions:

It can be stated that in this study and under the experimental conditions reported, the test item is not irritant to skin.

Executive summary:

The irritation potential of the test substance was investigated by using the in vitro Human Skin Model Test (OECD 439, EU B.46). Each three tissues of the human skin model EpiDerm were treated with 30 µL of the test item, the negative (Dulbecco's Phosphate Buffered Saline) or the positive control (5% sodium lauryl sulfate) for 60 minutes in total. Cell viability was determined using the MTT assay. The absorbance value obtained for the positive control was 0.081 and this result corresponds to 4.3% viability when compared to the results obtained from the negative controls. After treatment with the test item the mean relative absorbance value decreased irrelevantly to 99.9% compared to the relative absorbance value of the negative control. This value is not below the threshold for irritancy of ≤ 50%. Therefore, the test item is not considered to possess an irritant potential. Thus, the test substance is not considered to be irritant to skin under the test conditions chosen.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Eye irritation

Link to relevant study records

Reference

Endpoint:

eye irritation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2000-09-04 and 2000-09-08

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 405 (Acute Eye Irritation / Corrosion)

Version / remarks:

1987

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.5 (Acute Toxicity: Eye Irritation / Corrosion)

Version / remarks:

1997

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Species:

rabbit

Strain:

other: Little russian

Details on test animals or tissues and environmental conditions:

TEST ANIMALS
- Weight at study initiation: 1.6 — 1.8 kg bw
- Housing: individually
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: one week

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20 ± 3
- Humidity (%): 55 ± 15
- Air changes (per hr): 10
- Photoperiod (hrs dark / hrs light): 12/12

Vehicle:

unchanged (no vehicle)

Controls:

other: the untreated eye

Amount / concentration applied:

TEST MATERIAL
- Amount applied: 0.1 mL

Duration of treatment / exposure:

The lids were gently held together for 1 second.

Observation period (in vivo):

1 hour and 24, 48 and 72 hours

Number of animals or in vitro replicates:

4

Details on study design:

REMOVAL OF TEST SUBSTANCE
- Washing: none

SCORING SYSTEM: according to OECD scoring system

TOOL USED TO ASSESS SCORE: UV-light, fluorescein (at 24 hours)

Irritation parameter:

cornea opacity score

Basis:

animal #1

Time point:

24/48/72 h

Score:

0

Max. score:

4

Irritation parameter:

iris score

Basis:

animal #1

Time point:

24/48/72 h

Score:

0

Max. score:

2

Irritation parameter:

conjunctivae score

Basis:

animal #1

Time point:

24/48/72 h

Score:

0

Max. score:

3

Irritation parameter:

chemosis score

Basis:

animal #1

Time point:

24/48/72 h

Score:

0

Max. score:

4

Irritation parameter:

cornea opacity score

Basis:

animal #2

Time point:

24/48/72 h

Score:

0

Max. score:

4

Irritation parameter:

iris score

Basis:

animal #2

Time point:

24/48/72 h

Score:

0

Max. score:

2

Irritation parameter:

conjunctivae score

Basis:

animal #2

Time point:

24/48/72 h

Score:

0

Max. score:

3

Irritation parameter:

chemosis score

Basis:

animal #2

Time point:

24/48/72 h

Score:

0

Max. score:

4

Irritation parameter:

cornea opacity score

Basis:

animal #3

Time point:

24/48/72 h

Score:

0

Max. score:

4

Irritation parameter:

iris score

Basis:

animal #3

Time point:

24/48/72 h

Score:

0

Max. score:

2

Irritation parameter:

conjunctivae score

Basis:

animal #3

Time point:

24/48/72 h

Score:

0

Max. score:

3

Irritation parameter:

chemosis score

Basis:

animal #3

Time point:

24/48/72 h

Score:

0

Max. score:

4

Irritation parameter:

cornea opacity score

Basis:

animal #4

Time point:

24/48/72 h

Score:

0

Max. score:

4

Irritation parameter:

iris score

Basis:

animal #4

Time point:

24/48/72 h

Score:

0

Max. score:

2

Irritation parameter:

conjunctivae score

Basis:

animal #4

Time point:

24/48/72 h

Score:

0

Max. score:

3

Irritation parameter:

chemosis score

Basis:

animal #4

Time point:

24/48/72 h

Score:

0

Max. score:

4

Irritant / corrosive response data:

One hour after application of the test article animals No. 1, No. 2, No. 3 and No. 4 showed some conjuncitval vessels definitely injected, a swelling above normal and a discharge different from normal.
The cornea score was 0 for each individual animal before and after the instillation of Fluorescein.

Interpretation of results:

GHS criteria not met

Conclusions:

The test item is not considered to be irritating to rabbits eye.

Executive summary:

A study according to OECD 405 and EEC Guideline B.4 was performed with the test item to determine the irritating potential to rabbits eye. Four female albino rabbits were exposed to 0.1 mL of the test item in one eye. The eyes were examined and the changes were graded according to a numerical scale one hour, 24, 48 and 72 hours after dosing. One hour after application of the test item, all four animals showed some conjunctival vessels definitely injected, a swelling above normal an a discharge different from normal. 24 hours after application of the test article all four animals were free of any signs of eye irritation. 48 and 72 hours after application of the test article in none of the four animals were observed signs of eye irritation, too. The mean score (24/ 48/ 72 h) for cornea, iris, conjunctiva and chemosis were 0, 0, 0 and 0, respectively. In conclusion, the test item was not irritating to rabbits eye.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Respiratory irritation

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Skin Irritation

Key study

The irritation potential of the test substance was investigated by using the in vitro Human Skin Model Test (OECD 439, EU B.46). Each three tissues of the human skin model EpiDerm were treated with 30 µL of the test item, the negative (Dulbecco's Phosphate Buffered Saline) or the positive control (5% sodium lauryl sulfate) for 60 minutes in total. Cell viability was determined using the MTT assay. The absorbance value obtained for the positive control was 0.081 and this result corresponds to 4.3% viability when compared to the results obtained from the negative controls. After treatment with the test item the mean relative absorbance value decreased irrelevantly to 99.9% compared to the relative absorbance value of the negative control. This value is not below the threshold for irritancy of ≤ 50%. Therefore, the test item is not considered to possess an irritant potential. Thus, the test substance is not considered to be irritant to skin under the test conditions chosen.

The publication (Fuji et al. 1972) described two closed patch tests which were conducted on healthy male and female subjects. For the 48 hour closed patch test, 34 subjects were treated with 20% test substance in vaselinum aldum or unguentum hydrophilicum on the small back. In addition, a 24- to 72 -hour closed patch test was conducted on 30 male and female subjects, with the test item at 2% in unguentum simplex or unguentum hydrophilicum. In both tests, no irritant reactions were produced in the subjects (see section 7.10.5).

While no conclusion can be made of the irritation potential in the acute dermal toxicity study (Moreno, 1973), the study conduted with humans support the result of the key study.

Eye Irritation

A study according to OECD 405 and EEC Guideline B.4 was performed with the test item to determine the irritating potential to rabbits eye. Four female albino rabbits were exposed to 0.1 mL of the test item in one eye. The eyes were examined and the changes were graded according to a numerical scale one hour, 24, 48 and 72 hours after dosing. One hour after application of the test item, all four animals showed some conjunctival vessels definitely injected, a swelling above normal and a discharge different from normal. 24 hours after application of the test article all four animals were free of any signs of eye irritation. 48 and 72 hours after application of the test article in none of the four animals were observed signs of eye irritation, too. The mean (24, 48, 72 h) score for cornea, iris, conjunctiva and chemosis were 0, 0, 0 and 0, respectively. In conclusion, the test item was not irritating to rabbits eye.

Justification for classification or non-classification

Classification, Labelling, and Packaging Regulation (EC) No 1272/2008

Based on the available experimental data, the substance is not considered to be classified for skin or eye irritation under Regulation (EC) No 1272/2008,

as amended for the tenth time in Regulation (EC) No 2017/776.