12-oxahexadecan-16-olide

Endpoint summary

Description of key information

CERVOLIDE is not irritant in the in vitro skin irritation test under the experimental conditions described in this report.

In the in vivo study, CERVOLIDE was only observed at 24 hours after a single application and for up to 21 days following repeated application (days 1 to 21) at 3%, 10%, 30% and 100%.

When the in vitro experimental data is considered along side the in vivo study data, there is no evidence of corrosivity.

Therefore an in vitro corrosivity test is not required and CERVOLIDE will not be classified as a skin irritant.

The combination of the in vitro and in vivo tests indicate no eye irritation and therefore no classification is required.

Key value for chemical safety assessment

Skin irritation / corrosion

Link to relevant study records

Reference

Reference 1

Endpoint:

skin irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2017

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)

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

name of testing material : Cervolide
Batch:VE00445215
Expiry date: 07 June 2018

Test system:

human skin model

Source species:

human

Cell type:

non-transformed keratinocytes

Cell source:

other: SkinEthic Laboratories, Lyon, France.

Details on animal used as source of test system:

This model is a three-dimensional human epidermis model, which consists of adult human-derived epidermal keratinocytes which have been seeded on a dermal substitute consisting of a collagen type I matrix coated with type IV collagen. The keratinocytes were cultured for 13 days, which results in a highly differentiated and stratified epidermis model comprising the main basal, supra basal, spinous and granular layers and a functional stratum corneum.

Justification for test system used:

In the interest of sound science and animal welfare, a sequential testing strategy is recommended to minimize the need of in vivo testing. One of the validated in vitro skin irritation tests is the EPISKIN test, which is recommended in international guidelines (e.g. OECD and EC).

Vehicle:

unchanged (no vehicle)

Details on test system:

On the day of receipt the tissues were transferred to 12-well plates and pre-incubated with pre-warmed Maintenance Medium for 3.5 hours at 37°C. Maintenance medium and Assay medium were supplied by Skinethic Laboratories, Lyon, France.MTT concentrate (Sigma Aldrich, Zwijndrecht, The Netherlands; 3 mg/ml in PBS) diluted (10x) in Assay medium (final concentration 0.3 mg/ml).

Control samples:

yes, concurrent negative control

yes, concurrent positive control

Amount/concentration applied:

undiluted (25 µl) directly

Duration of treatment / exposure:

15 ± 0.5 minutes

Duration of post-treatment incubation (if applicable):

After a 42 hour post-incubation period, determination of the cytotoxic (irritancy) effect was performed. Cytotoxicity is expressed as the reduction of mitochondrial dehydrogenase activity measured by formazan production from MTT at the end of the treatment.

Irritation / corrosion parameter:

% tissue viability

Value:

83

Vehicle controls validity:

valid

Negative controls validity:

valid

Positive controls validity:

valid

The positive control had a mean cell viability of 8.8% after 15 ± 0.5 minutes exposure. The absolute mean OD₅₇₀(optical density at 570 nm) of the negative control tissues was within the laboratory historical control data range. The standard deviation value of the percentage viability of three tissues treated identically was less than 17%, indicating that the test system functioned properly.

Skin irritation is expressed as the remaining cell viability after exposure to the test item. The relative mean tissue viability obtained after 15 ± 0.5 minutes treatment with CERVOLIDE compared to the negative control tissues was 83%. Since the mean relative tissue viability for CERVOLIDE was above 50% after 15 ± 0.5 minutes treatment CERVOLIDE is considered to be non-irritant.

In conclusion, CERVOLIDE is non-irritant in the in vitro skin irritation test under the experimental conditions described in this report.

Interpretation of results:

GHS criteria not met

Conclusions:

The relative mean tissue viability obtained after 15 ± 0.5 minutes treatment with CERVOLIDE compared to the negative control tissues was 83%. Since the mean relative tissue viability for CERVOLIDE was above 50% after 15 ± 0.5 minutes treatment CERVOLIDE is considered to be non-irritant.

Executive summary:

The objective of this study was to evaluate CERVOLIDE for its ability to induce skin irritation on a human three dimensional epidermal model (EPISKIN Small model (EPISKIN-SM^TM)). The possible skin irritation potential of CERVOLIDE was tested through topical application for 15 minutes.  The study procedures described in this report were based on the most recent OECD and EC guidelines.

The relative mean tissue viability obtained after 15 ± 0.5 minutes treatment with CERVOLIDE compared to the negative control tissues was 83%. Since the mean relative tissue viability for CERVOLIDE was above 50% after 15 ± 0.5 minutes treatment CERVOLIDE is considered to be non-irritant.

In conclusion, CERVOLIDE is non-irritant in the in vitro skin irritation test under the experimental conditions described in this report.

Reference 2

Endpoint:

skin irritation: in vivo

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

1977

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Qualifier:

according to guideline

Guideline:

other: Skin irritation and capacity of allergenic sensitization determined by Open Epicutaneous test on Guinea Pigs

Principles of method if other than guideline:

Single application to assess irritantcy prior to induction procedure and also after multiple application of the test substance up to 21 days.

GLP compliance:

no

Specific details on test material used for the study:

Test material name (as stated in the report): Giv 81-1543

Species:

guinea pig

Strain:

not specified

Details on test animals or test system and environmental conditions:

One to six experimental groups and one control group of 6 to 8 guinea pigs were used.

Type of coverage:

open

Preparation of test site:

clipped

Vehicle:

other: either ethanol, acetone, H2O, vasoline or PEG.

Controls:

yes, concurrent no treatment

Amount / concentration applied:

Single application: 0.025 ml of undiluted test material and at100%, 30%, 10%, and 3%.
Multiple application: 0.1 ml of test material at 100%, 30%, 10%, and 3% for 21 days.

Duration of treatment / exposure:

Single application
Multiple application for 21 days

Observation period:

Single day and up to 21 days.

Number of animals:

6 to 8 guinea pigs were used per group.

Details on study design:

One day before starting the induction procedure the threshold toxic concentration of CERVOLIDE was estimated.
A single application of 0.025mL of each concentration is simultaneously performed on one of the areas measuring 2 cm2 of the flank skin previously clipped and marked with a circular stamp.
The skin reactions were read 24 hours after the application of the test material.
The minimal irritant and the non-irritant concentrations were determined by an all or none criteria.

Induction - Application of 0.1 ml for the OET procedure.

Repeated application of Cetonal on days 1 to 21 were also performed with reactions noted on days 7, 14 and 21.

Irritation parameter:

overall irritation score

Basis:

mean

Time point:

7 d

Score:

4

Max. score:

4

Reversibility:

not reversible

Irritation parameter:

overall irritation score

Basis:

mean

Time point:

14 d

Score:

4

Max. score:

4

Reversibility:

not reversible

Irritation parameter:

overall irritation score

Basis:

mean

Time point:

21 d

Score:

4

Max. score:

4

Reversibility:

not reversible

Irritation parameter:

overall irritation score

Basis:

mean

Time point:

24 h

Score:

4

Max. score:

4

Reversibility:

other: only 24h conducted

Remarks on result:

positive indication of irritation

Interpretation of results:

Category 3 (mild irritant) based on GHS criteria

Conclusions:

Reactions were evaluated at the end of each week.
Very slight irritation was observed with 3% after 7, 14 and 21 days. With 10%, very slight irritation was observed after 7 and 14 days, with slight irritation after 21 days. With 30%, slight irritation was observed
after 7 days, with moderate irritation after 14 and 21 days. With the neat material, moderate irritation was observed after 7 and 14 days, with strong irritation after 21 days.

Executive summary:

The CERVOLIDE was tested in this study by Open Epicutaneous test on Guinea Pigs Pre-Guidance but according to the practice devised by Klecak at the time.

One day before starting the induction procedure the threshold toxic concentration of CERVOLIDE was estimated.

A single application of 0.025mL of each concentration is simultaneously performed on one of the areas measuring 2 cm2 of the flank skin previously clipped and marked with a circular stamp.

The skin reactions were read 24 hours after the application of the test material.

The minimal irritant and the non-irritant concentrations were determined by an all or none criteria.

Induction - Application of 0.1 ml for the OET procedure.

Repeated application of CERVOLIDE on days 1 to 21 were also performed with reactions noted on days 7, 14 and 21.

Very slight irritation was observed with 3% after 7, 14 and 21 days. With 10%, very slight irritation was observed after 7 and 14 days, with slight irritation after 21 days. With 30%, slight irritation was observed

after 7 days, with moderate irritation after 14 and 21 days. With the neat material, moderate irritation was observed after 7 and 14 days, with strong irritation after 21 days.

According to this test, Cetronal is clasified Category 3 (mild irritant) based on GHS criteria.

Reference 3

Endpoint:

skin irritation: in vivo

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

1977

Reliability:

4 (not assignable)

Rationale for reliability incl. deficiencies:

abstract

GLP compliance:

no

Specific details on test material used for the study:

Name of testing material: 77-227 Hibiscolide

Species:

rabbit

Strain:

not specified

Irritation parameter:

overall irritation score

Basis:

mean

Time point:

24 h

Score:

4

Max. score:

4

Reversibility:

not reversible

Interpretation of results:

Category 2 (irritant) based on GHS criteria

Conclusions:

As part of a acute dermal LD50 study, 10 rabbits were observed for dermal reactions. Irritation was evaluated on day 1.
Irritation was evaluated during a dermal LD50 study, in which 10 rabbits were treated with 5000 mg/kg of 12-oxahexadecanolide. Mild (9/10) to moderate (1/10) erythema and mild edema (1/10) was observed

Executive summary:

As part of a acute dermal LD50 study, 10 rabbits were observed for dermal reactions. Irritation was evaluated on day 1.

Irritation was evaluated during a dermal LD50 study, in which 10 rabbits were treated with 5000 mg/kg of 12-oxahexadecanolide. Mild (9/10) to moderate (1/10) erythema and mild edema (1/10) was observed

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:

1977

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Qualifier:

according to guideline

Guideline:

other: The method of procedure is that suggested by Dr. Draize and described in "Appraisal of the safety of chemicals in foods, drugs amd cosmetics" published by the Association of Food and Drug Officials of the United States.

GLP compliance:

no

Specific details on test material used for the study:

Test material name (as stated in the report): GIV 81-1543=Musc 174

Species:

rabbit

Strain:

other: albino rabbits

Details on test animals or tissues and environmental conditions:

Three normal, healthy albino rabbit were used in this experiment

Controls:

yes, concurrent no treatment

Amount / concentration applied:

0.1mL of test materia

Duration of treatment / exposure:

instillation

Observation period (in vivo):

14 days after instillation

Number of animals or in vitro replicates:

3 rabbits

Details on study design:

Ecah animal had 0.1 mL of test sample instilled into the right eye with no further treatment.

Irritation parameter:

cornea opacity score

Basis:

animal:

Remarks:

animal #1 ; animal #2 and animal 3#

Time point:

24/48/72 h

Score:

0

Max. score:

0

Reversibility:

other: Na

Irritation parameter:

iris score

Basis:

animal: animal #1 ; animal #2 and animal 3#

Time point:

24/48/72 h

Score:

0

Max. score:

0

Reversibility:

other: Na

Irritation parameter:

conjunctivae score

Basis:

animal: animal #1 ; animal #2 and animal 3#

Time point:

24/48/72 h

Score:

0

Max. score:

1

Reversibility:

fully reversible within: 72h

Irritation parameter:

chemosis score

Basis:

animal: animal #1 ; animal #2 and animal 3#

Time point:

24/48/72 h

Score:

0

Max. score:

0

Reversibility:

other: Na

Irritation parameter:

other: Ulceration

Basis:

animal: animal #1 ; animal #2 and animal 3#

Time point:

24/48/72 h

Score:

0

Max. score:

0

Reversibility:

other: Na

The scorings recorded were made according to the Draize scale for scoring ocular lesions.

Interpretation of results:

GHS criteria not met

Conclusions:

CERVOLIDE does not meet the criteria to be as Eye irritant Category 2  according to the CLP Regulation (EC) No. 1272/2008.

Executive summary:

The CERVOLIDE was tested in this study to determine his eye irritant potential according to the procedure is that suggested by Dr. Draize and described in "Appraisal of the safety of chemicals in foods, drugs amd cosmetics" published by the Association of Food and Drug Officials of the United States.

Observations and scoring by the Draize method were conducted immediately and 1 hour, 24 hours, 48 hours, 72 hours, 7 days and 14 days later. With 10%, conjunctival redness was observed in 3/3 animals at 1 hour and 24 hours. Reactions were absent by 48 hours. With 30%, conjunctival redness was observed in 3/3, 3/3 and 1/3 animals at 1 hour, 24 hours and 48 hours, respectively. Reactions were absent by 72 hours.With the neat material, conjunctival redness was observed in 3/3, 1/3 and 3/3 animals at 1 hour, 24 hours and 48 hours, respectively. Moderate conjunctival redness was observed in 2/3 animals at 24 hours. Chemosis was observed in 3/3 animals at 24 hours. Dose: 100 % Effects: eye effects; irritant effects Results: moderate conjunctival irritation which cleared within 72 hours

CERVOLIDE does not meet the criteria to be as Eye irritant Category 2  according to the CLP Regulation (EC) No. 1272/2008.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Justification for classification or non-classification

When the in vitro experimental data is considered alongside the in vivo study data, there is no evidence of corrosivity.

Therefore an in vitro corrosivity test is not required and CERVOLIDE will not be classified as a skin irritant.

The combination of the in vitro and in vivo tests indicate no eye irritation and therefore no classification is required.