Reaction mass of undec-8-enal and undec-9-enal and undec-10-enal

Endpoint summary

Description of key information

Skin corrosion: Not corrosive because the substance is not irritating to skin and eyes.

Skin irritation (OECD TG 439): not irritating

Eye irritation (OECD TG 438): not irritating

Respiratory irritation: no adverse effects in absence of: human data indicating such, corrosion or irritation

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:

04-01-2017 to 10-02-2017

Reliability:

1 (reliable without restriction)

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

July 28, 2015

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

No 761/2009

Qualifier:

according to guideline

Guideline:

other: UN GHS

Version / remarks:

published 2003, last (6th) revision 2015

GLP compliance:

yes (incl. QA statement)

Test system:

human skin model

Source species:

human

Cell type:

non-transformed keratinocytes

Cell source:

other: SkinEthic Laboratories (Lyon, France)

Source strain:

other: not applicable

Justification for test system used:

According to testing guideline OECD Guideline 439

Vehicle:

unchanged (no vehicle)

Details on test system:

RECONSTRUCTED HUMAN EPIDERMIS (RHE) TISSUE
- Model used: EPISKIN TM
- Tissue batch number(s): 17-EKIN-006
- Production date/Shipping date: no data
- Delivery date: 07 February 2017
- Date of initiation of testing: 07 February 2017

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

REMOVAL OF TEST MATERIAL AND CONTROLS
-Tissues were washed with phosphate buffered saline to remove residual test item.

MTT DYE USED TO MEASURE TISSUE VIABILITY AFTER TREATMENT / EXPOSURE
- MTT concentration: 0.3 mg/ml in PBS
- Incubation time: 3 hours at 37 °C
- Spectrophotometer: not specified
- Wavelength: 570 nm
- Filter: no information

NUMBER OF REPLICATE TISSUES: 3

DECISION CRITERIA
- A test substance is considered irritant in the skin irritation test if: The relative mean tissue viability of three individual tissues after 15 minutes of exposure to the test substance and 42 hours of post incubation is ≤ 50% of the mean viability of the negative controls.
- A test substance is considered non-irritant in the in vitro skin irritation test if: The relative mean tissue viability of three individual tissues after 15 minutes of exposure to the test substance and 42 hours of post incubation is > 50% of the mean viability of the negative controls.

Control samples:

yes, concurrent negative control

yes, concurrent positive control

Amount/concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 10 μL

Duration of treatment / exposure:

15 minutes

Duration of post-treatment incubation (if applicable):

42 hours

Number of replicates:

3

Irritation / corrosion parameter:

% tissue viability

Remarks:

Relative mean tissue viability compared to the negative control tissues (100%)

Value:

52.9

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Remarks:

9.7%

Remarks on result:

no indication of irritation

Other effects / acceptance of results:

OTHER EFFECTS:
- Direct-MTT reduction: No colour changes observed
- Colour interference with MTT: No colour changes observed

ACCEPTANCE OF RESULTS:
-The relative mean tissue viability for the positive control treated tissues was 9.7% relative to the negative control treated tissues and the standard deviation value of the viability was 13.8%. The positive control acceptance criteria were therefore satisfied.
-The mean OD570 for the negative control treated tissues was between ≥ 0.6 and ≤ 1.5 and the standard deviation value of the viability was 13.9%. The negative control acceptance criteria were therefore satisfied.
-The standard deviation calculated from individual tissue viabilities of the three identically test item treated tissues was 14.7%. The test item acceptance criterion was therefore satisfied.

Results after treatment with Intreleven Aldehyde and controls

Test Group	Mean Absorbance of 3 Tissues	Relative Absorbance [%] Tissue 1, 2 + 3**	Relative Standard Deviation [%]	Rel. Absorbance [% of Negative Control]***
Negative Control	0.888	101.0 / 113.4 / 85.6	13.9	100.0
Positive Control	0.086	8.4 / 9.6 / 11.1	13.8	9.7
Test Item	0.470	61.8 / 47.4 / 49.5	14.7	52.9

* Mean of two replicate wells after blank correction

** relative absorbance per tissue [rounded values]

*** relative absorbance per treatment group [rounded values]

Interpretation of results:

other: Not skin irritant

Remarks:

according to EU CLP criteria (1272/2008/EC and its updates)

Conclusions:

Under the conditions of this test, the relative mean tissue viability for the test item was determined as 52.9%, therefore Intreleven aldehyde is not a skin irritant.

Executive summary:

The possible skin irritation potential of Intreleven aldehyde was tested in vitro using a human skin model through topical application. The study procedures described in this report were according to OECD TG 439 guideline and GLP principles. Skin tissue was treated by topical application of 10 μL undiluted test substance for 15 minutes. After further 42 hours 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 (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl-tetrazolium bromide) MTT at the end of treatment. Skin irritation is expressed as the remaining cell viability after exposure to the test substance. Reliable negative and positive controls were included. The positive control had a mean cell viability of 9.7% after 15 minutes exposure. The standard deviation value of the percentage viability of three tissues treated identically was less than 18%, indicating that the test system functioned properly. Under the conditions of this test, the relative mean tissue viability for the test item was determined to be 52.9%. This value is above the threshold for irritancy of ≤50%, therefore Intreleven aldehyde is not a skin irritant.

Reference 2

Endpoint:

skin irritation: in vitro / ex vivo

Data waiving:

study scientifically not necessary / other information available

Justification for data waiving:

other:

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:

23-12-2016 to 08-02-2017

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 438 (Isolated Chicken Eye Test Method for Identifying i) Chemicals Inducing Serious Eye Damage and ii) Chemicals Not Requiring Classification for Eye Irritation or Serious Eye Damage)

GLP compliance:

yes (incl. QA statement)

Remarks:

Triskelion B.V., Utrechtseweg 48, 3700 AV, Zeist

Species:

other: Eyes of male or female chickens (ROSS, spring chickens)

Details on test animals or tissues and environmental conditions:

SOURCE OF COLLECTED EYES
- Source: Slaughterhouse v.d. Bor, Nijkerkerveen, The Netherlands
- Characteristics of donor animals: Approximately 7 weeks old, male or female chickens, body weight range approximately 1.5-2.5 kg, were used as eye donors.
- Storage, temperature and transport conditions of ocular tissue: Heads of the animals were cut off immediately after sedation of the animals by electric shock and incision of the neck for bleeding, and before they reached the next station on the process line. The heads were placed in small plastic boxes on a bedding of paper tissues moistened with isotonic saline. Next, they were transported to the testing facility. During transportation, the heads were kept at ambient temperature.
- Time interval prior to initiating testing: Within 2 hours after kill, eyes were carefully dissected and placed in a superfusion apparatus.
- Indication of any existing defects or lesions in ocular tissue samples: No
- Indication of any antibiotics used: No

Vehicle:

unchanged (no vehicle)

Details on study design:

SELECTION AND PREPARATION OF ISOLATED EYES
Within 2 hours after kill, eyes were carefully dissected and placed in a superfusion apparatus using the following procedure: First the eye-lids were carefully removed without damaging the cornea and a small drop of Fluorescein sodium 2.0% w/v was applied to the corneal surface for a few seconds and subsequently rinsed off with isotonic saline at ambient temperature. Next, the head with the fluorescein-treated cornea was examined with a slit-lamp microscope (Slit-lamp 900 BP, Haag-Streit AG, Liebefeld-Bern, Switzerland) to ensure that the cornea was not damaged. If undamaged (e.g., fluorescein retention and corneal opacity scores of ≤ 0.5), the eye was further dissected from the head without damaging the eye or cornea. Care was taken to remove the eye-ball from the orbit without cutting off the optical nerve too short. The enucleated eye was placed in a stainless steel clamp with the cornea positioned vertically and transferred to a chamber of the superfusion apparatus. The clamp holding the eye was positioned in such a way that the entire cornea was supplied with isotonic saline from a bent, stainless steel tube, at a target rate of 0.10-0.15 mL/min. The chambers of the superfusion apparatus as well as the saline were temperature controlled at approximately 32 °C (water pump set at 36.4 °C). After placing in the superfusion apparatus, the eyes were examined again with the slit-lamp microscope to ensure that they were not damaged. An accurate measurement was taken at the corneal apex of each eye. Eyes with a corneal thickness deviating more than 10% of the average corneal thickness of the eyes, eyes showing opacity (score higher than 0.5), or were unacceptably stained with fluorescein (score higher than 0.5) indicating the cornea to be permeable, or eyes that showed any other signs of damage, were rejected as test eyes and replaced.

EQUILIBRATION AND BASELINE RECORDINGS
Each eye provided its own baseline values for corneal swelling, corneal opacity and fluorescein retention. For that purpose, after an equilibration period of 45-60 minutes, the corneal thickness of the eyes was measured again to determine the zero reference value for corneal swelling calculations.

NUMBER OF REPLICATES
Negative control: 1
Positive control: 3
Test group: 3

NEGATIVE CONTROL USED
Physiological saline

POSITIVE CONTROL USED
Benzalkonium Chloride 5%

APPLICATION DOSE AND EXPOSURE TIME
30 μL for 10 seconds

OBSERVATION PERIOD
240 minutes

REMOVAL OF TEST SUBSTANCE
- Volume and washing procedure after exposure period: 20 mL saline. After rinsing, each eye in the holder was returned to its chamber.
- Indicate any deviation from test procedure in the Guideline: none

METHODS FOR MEASURED ENDPOINTS:
- Corneal opacity: Slit-lamp microscope examination
- Damage to epithelium based on fluorescein retention: Slit-lamp microscope examination
- Swelling: measured with optical pachymeter on a slit-lamp microscope; slit-width setting: set at 0.095 mm
- Others: After the final examination, the test substance treated eyes, the negative and positive control eyes were preserved in a neutral aqueous phosphate-buffered 4% solution of formaldehyde. The corneas were embedded in paraffin wax, sectioned at ca 4 μm and stained with PAS (Periodic Acid-Schiff). The microscopic slides were subjected to histopathological examination.

SCORING SYSTEM:
Defined scoring scales were used for each parameter to define the severity of effects into four categories (I-IV).
- Mean corneal swelling (%): According to OECD 438 guideline. Examination of the eyes after 0, 30, 75, 120, 180, and 240 minutes
- Mean maximum opacity score: According to OECD 438 guideline. Examination of the eyes after 0, 30, 75, 120, 180, and 240 minutes
- Mean fluorescein retention score at 30 minutes post-treatment: According to OECD 438 guideline.

DECISION CRITERIA: According to OECD 438 guideline

Irritation parameter:

percent corneal swelling

Run / experiment:

slit-lamp examination

Value:

0

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

other: maximum mean values

Irritation parameter:

cornea opacity score

Run / experiment:

slit-lamp examination

Value:

0

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

other: maximum mean values

Irritation parameter:

fluorescein retention score

Run / experiment:

slit-lamp examination

Value:

0

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Other effects / acceptance of results:

Slit-lamp examination: the test substance did not cause any corneal effects. The negative control eye did not show any corneal effect and demonstrated that the general conditions during the tests were adequate. The positive control BAC 5% caused severe corneal effects and demonstrated the ICE test valid to detect severe eye irritants.

Microscopic examination of the corneas treated with the test substance did not reveal any abnormalities other than very slight erosion of the epithelium in one cornea, which is considered an acceptable background finding. Microscopic examination of the cornea treated with the negative control (saline) did not reveal any abnormalities. Microscopic examination of the corneas treated with the positive control BAC 5% revealed slight, moderate or severe erosion and very slight or slight vacuolation of the epithelium and the epithelium partly detached from the basement membrane.

Interpretation of results:

other: Not eye irritant

Remarks:

according to EU CLP criteria (1272/2008/EC and its updates)

Conclusions:

Under the test conditions (OECD 438 and GLP) the test substance is not considered to be an eye irritant.

Executive summary:

In accordance with OECD guideline 438 and GLP, the test substance was examined in vitro for its eye irritating potential using the Isolated Chicken Eye (ICE) Test. In this test, 3 eyes were exposed to 30 µL test substance for 10 seconds. In addition, one negative control eye (30 µL saline) and three positive control eyes (30 µL Benzalkonium Chloride (BAC)) were tested. After the exposure the eyes were rinsed with 20 mL saline and were examined at approximately 0, 30, 75, 120, 180, and 240 minutes after treatment. The test substance did not cause any corneal effects. The negative control eye did not show any corneal effect and demonstrated that the general conditions during the test were adequate. The positive control BAC 5% caused severe corneal effects and demonstrated the ICE test valid to detect severe eye irritants. Microscopic examination of the corneas treated with the test substance did not reveal any abnormalities other than very slight erosion of the epithelium in one cornea, which is considered an acceptable background finding. Microscopic examination of the cornea treated with the negative control (saline) did not reveal any abnormalities. Microscopic examination of the corneas treated with the positive control BAC 5% revealed slight, moderate or severe erosion and very slight or slight vacuolation of the epithelium and the epithelium partly detached from the basement membrane. Based on these results, the test substance is not considered to be an eye irritant.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Respiratory irritation

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Additional information

Skin corrosion

The substance is not corrosive because the substance is not irritating to skin and eyes.

Skin irritation

The possible skin irritation potential of Intreleven aldehyde was tested in vitro using a human skin model through topical application. The study procedures described in this report were according to OECD TG 439 guideline and GLP principles. Skin tissue was treated by topical application of 10 μL undiluted test substance for 15 minutes. After further 42 hours 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 (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl-tetrazolium bromide) MTT at the end of treatment. Skin irritation is expressed as the remaining cell viability after exposure to the test substance. Reliable negative and positive controls were included. The positive control had a mean cell viability of 9.7% after 15 minutes exposure. The standard deviation value of the percentage viability of three tissues treated identically was less than 18%, indicating that the test system functioned properly. Under the conditions of this test, the relative mean tissue viability for the test item was determined to be 52.9%. This value is above the threshold for irritancy of ≤50%, therefore Intreleven aldehyde is not a skin irritant.

 

Eye irritation

In accordance with OECD guideline 438 and GLP, the test substance was examined in vitro for its eye irritating potential using the Isolated Chicken Eye (ICE) Test. In this test, 3 eyes were exposed to 30 µL test substance for 10 seconds. In addition, one negative control eye (30 µL saline) and three positive control eyes (30 µL Benzalkonium Chloride (BAC)) were tested. After the exposure the eyes were rinsed with 20 mL saline and were examined at approximately 0, 30, 75, 120, 180, and 240 minutes after treatment. The test substance did not cause any corneal effects. The negative control eye did not show any corneal effect and demonstrated that the general conditions during the test were adequate. The positive control BAC 5% caused severe corneal effects and demonstrated the ICE test valid to detect severe eye irritants. Microscopic examination of the corneas treated with the test substance did not reveal any abnormalities other than very slight erosion of the epithelium in one cornea, which is considered an acceptable background finding. Microscopic examination of the cornea treated with the negative control (saline) did not reveal any abnormalities. Microscopic examination of the corneas treated with the positive control BAC 5% revealed slight, moderate or severe erosion and very slight or slight vacuolation of the epithelium and the epithelium partly detached from the basement membrane. Based on these results, the test substance is not considered to be an eye irritant.

 

Respiratory irritation

Respiratory irritation is not anticipated because there are no human data indicating such effects. In addition, the substance is not a skin or eye irritant under in vitro conditions, which further supports the absence of concern for respiratory irritation.

Justification for classification or non-classification

Based on the available data, Intreleven aldehyde was not classified for skin, eye or respiratory irritation in accordance with the criteria outlined in the EU CLP Regulation (EC No. 1272/2008 and its updates).