Registration Dossier

Diss Factsheets

Administrative data

Description of key information

Skin corrosion: the substance is not corrosive based on the absence of acidic, base or other very reactive groups

Skin irritation: irritating based on read across from Myrcenyl acetate 'mono', tested in OECD TG 439

Eye irritation: not eye irritating based on Myrcenyl acetate 'mono' tested in OECD TG 438

Key value for chemical safety assessment

Skin irritation / corrosion

Link to relevant study records

Referenceopen allclose all

Endpoint:
skin irritation: in vitro / ex vivo
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Study period:
2018
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Read-across information.
Justification for type of information:
The read across rationale is presented in the related Endpoint summary, the accompanying files are also attached there.
Reason / purpose for cross-reference:
read-across source
Irritation / corrosion parameter:
other: Relative mean viability
Value:
16.2
Negative controls validity:
valid
Positive controls validity:
valid
Interpretation of results:
other: Skin irritant Category 2
Remarks:
according to EU CLP (EC No. 1272/2008 and its amendments).
Conclusions:
For the substance a relative viability of 16.2% is derived (< 50%) based on read across from Myrcenyl acetate 'mono'.
Endpoint:
skin irritation: in vitro / ex vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
Experimental start date 24 May 2016, experimental completion date 30 May 2016
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Justification for type of information:
The information is used for read across to Pseudo linalyl acetate.
Qualifier:
according to guideline
Guideline:
OECD Guideline 439 (In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method)
Version / remarks:
Adopted 28 July 2015
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Test system:
other: EPISKIN reconstructed human epidermis model
Source species:
human
Cell type:
other: adult human-derived epidermal keratinocytes
Source strain:
other: Not applicable
Vehicle:
unchanged (no vehicle)
Details on test system:
TEST SYSTEM:
EPISKIN™ Reconstructed Human Epidermis Model Kit:
Supplier: SkinEthic Laboratories, Lyon, France
Date received: 24 May 2016
EpiSkinTM Tissues (0.38cm2) lot number: 16-EKIN-021
Maintenance Medium lot number: 16-MAIN3-035
Assay Medium lot number: 16-ESSC-021

The EPISKIN model is a three-dimensional reconstructed human epidermis model consisting of adult human-derived epidermal keratinocytes seeded on a dermal substitute consisting of a collagen type I matrix coated with type IV collagen. A highly differentiated and stratified epidermis model is obtained after a 13-Day culture period comprising of the main basal, supra basal, spinous and granular layers and a functional stratum corneum.

PRE-TEST PROCEDURE:
Assessment of Direct Test Item Reduction of MTT:
MTT Salt Metabolism, Cell Viability Assay:
The MTT assay, a colorimetric method of determining cell viability, is based on reduction of the yellow tetrazolium salt (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl-tetrazolium bromide) to a blue formazan salt by mitochondrial succinate dehydrogenase in viable cells.

One limitation of the assay is possible interference of the test item with MTT. A test item may directly reduce MTT, thus mimicking dehydrogenase activity of the cellular mitochondria. This property of the test item is only a problem, if at the time of the MTT test (after rinsing) there are still sufficient amounts of the test item present on or in the tissues. In this case, the true metabolic MTT reduction and the false direct MTT reduction can be differentiated and quantified by using killed tissues to act as controls.

Test for Direct MTT Reduction:
As specified, a test item may interfere with the MTT endpoint, if it is able to directly reduce MTT and at the same time is present on or in the tissues when the MTT viability test is performed. To identify this possible interference, the test item is checked for the ability to directly reduce MTT according to the following procedure:

10 µL of the test item was added to 2 mL of a 0.3 mg/mL MTT solution freshly prepared in assay medium. The solution was incubated in the dark at 37 °C, 5% CO2 in air for 3 hours. Untreated MTT solution was used as a control.

If the MTT solution containing the test item turns blue or purple, the test item is presumed to have reduced the MTT and the determination of skin irritation potential would be performed in parallel on viable and water-killed tissues for quantitative correction of the results.

Assessment of Color Interference with the MTT endpoint:
A test item may interfere with the MTT endpoint if it is colored. The MTT assay is affected only if the test item is present in the tissues when the MTT viability assay is performed.

10 µL of test item was added to 90 µL of sterile water. After mixing for 15 minutes on a plate shaker a visual assessment of the color was made.

PRE-INCUBATION (Day 0: TISSUE ARRIVAL):
Before removal from the transport plate each tissue was inspected for any air bubbles between the agarose gel and the insert:
Tissues Satisfactory: Yes
Temperature Indicator Color Satisfactory: Yes
Agar Medium Color Satisfactory: Yes

2 mL of maintenance medium, warmed to approximately 37 °C, was pipetted into the first column of 3 wells of a pre-labeled 12-well plate. Each epidermis unit was transferred into the maintenance medium filled wells (3 units per plate). A different 12-well plate was used for the test item and each control item. The tissues were incubated at 37 °C, 5% CO2 in air overnight.

MAIN TEST:
Application of Test Item and Rinsing (Day 1):
2 mL of maintenance medium, warmed to approximately 37 °C, was pipetted into the second column of 3 wells of the 12 well plate.

Triplicate tissues were treated with the test item for an exposure period of 15 minutes. The test item was applied topically to the corresponding tissues ensuring uniform covering. 10 µL (26.3 µL/cm2) of the test item was applied to the epidermis surface. Triplicate tissues treated with 10 µL of DPBS served as the negative controls and triplicate tissues treated with 10 µL of SDS 5% w/v served as the positive controls. To ensure satisfactory contact with the positive control item the SDS solution was spread over the entire surface of the epidermis using a pipette tip (taking particular care to cover the center). After a 7-Minute contact time the SDS solution was re-spread with a pipette tip to maintain the distribution of the SDS for the remainder of the contact period (re-spreading is not required for the negative control or test item). The plates were kept in the biological safety cabinet at room temperature for 15 minutes.

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. The rinsed tissues were transferred to the second column of 3 wells containing 2 mL of maintenance medium in each well. The rinsed tissues were incubated at 37 °C, 5% CO2 in air for 42 hours.

MTT Loading/Formazan Extraction (Day 3):
Following the 42-Hour post-exposure incubation period each 12-well plate was placed onto a plate shaker for 15 minutes to homogenize the released mediators in the maintenance medium. 1.6 mL of the maintenance medium from beneath each tissue was transferred to pre-labeled micro tubes and stored in a freezer at -14 to -30 ºC for possible inflammatory mediator determination.

2 mL of a 0.3 mg/mL MTT solution, freshly prepared in assay medium, was pipetted into the third column of 3 wells of the 12-well plates. The tissues were transferred to the MTT filled wells, being careful to remove any excess maintenance medium from the bottom of the tissue insert by blotting on absorbent paper. The tissues were incubated for 3 hours at 37 °C, 5% CO2 in air. At the end of the 3-Hour incubation period each tissue was placed onto absorbent paper to dry. A total biopsy of the epidermis was made using the EPISKIN biopsy punch. The epidermis was carefully separated from the collagen matrix using forceps and both parts (epidermis and collagen matrix) placed into labeled 1.5 mL micro tubes containing 500 µL of acidified isopropanol, ensuring that both the epidermis and collagen matrix were fully immersed. Each tube was plugged to prevent evaporation and mixed thoroughly on a vortex mixer. The tubes were refrigerated at 1 to 10 °C until Day 6 of the experiment, allowing the extraction of formazan crystals out of the MTT-loaded tissues.

Absorbance/Optical Density Measurements (Day 6):
At the end of the formazan extraction period each tube was mixed thoroughly on a vortex mixer to produce a homogenous colored solution.

For each tissue, duplicate 200 µL samples were transferred to the appropriate wells of a pre-labeled 96-well plate. 200 µL of acidified isopropanol alone was added to the two wells designated as ‘blanks’. The optical density was measured (quantitative viability analysis) at 562 nm (without a reference filter) using the Anthos 2001 microplate reader.

DATA EVALUATION:
Quantitative MTT Assessment (Percentage Tissue Viability):
For the test item the mean tissue viability obtained after the 15-Minute exposure period followed by the 42-Hour post-exposure incubation period were compared to the mean of the negative control treated tissues (n=3) to generate the relative mean tissue viability. The relative mean viability was calculated in the following way:
Relative mean viability (%) = ((mean OD562 of test item) / (mean OD562 of negative control)) * 100

Classification of irritation potential is based upon relative mean tissue viability following the 15-Minute exposure period followed by the 42-Hour post-exposure incubation period according to the following table:

Criteria for in vitro interpretation: Prediction
Relative mean tissue viability is ≤50%: Irritant
Relative mean tissue viability is >50%: Non-irritant

Quality Criteria:
The results of the assay are considered acceptable if the following assay acceptance criteria are achieved:
Positive Control:
The assay establishes the acceptance criterion for an acceptable test if the relative mean tissue viability for the positive control treated tissues is ≤40% relative to the negative control treated tissues, and the standard deviation (SD) value of the percentage viability is ≤18%.

Negative Control:
The assay establishes the acceptance criterion for an acceptable test if the mean OD562 for the negative control treated tissues is ≥0.6 and ≤1.5, and the SD value of the percentage viability is ≤18%.

Test Item:
The assay establishes the acceptance criterion for an acceptable test if the standard deviation calculated from individual percentage tissue viabilities of the three identically treated tissues is ≤18%.

Major Computerized Systems:
The following computerized system was used in the study:
Delta Building Monitoring System
Control samples:
yes, concurrent negative control
yes, concurrent positive control
Amount/concentration applied:
10 µL
Duration of treatment / exposure:
15 minutes
Duration of post-treatment incubation (if applicable):
42 hours
Number of replicates:
3
Irritation / corrosion parameter:
other: Relative mean viability (%)
Value:
16.2
Vehicle controls validity:
not applicable
Negative controls validity:
valid
Positive controls validity:
valid
Other effects / acceptance of results:
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.

Assessment of Color Interference with the MTT endpoint:
The solution (10 µL of test item in 90 µL of sterile water) containing the test item was colorless. It was therefore unnecessary to run color correction tissues.

Test Item, Positive Control Item and Negative Control Item:
The individual and mean OD562 values, standard deviations and tissue viabilities for the test item, negative control item and positive control item are given under "Any other information on results incl. tables". The mean viabilities and standard deviations of the test item and positive control, relative to the negative control are also given under "Any other information on results incl. tables".

The relative mean viability of the test item treated tissues was 16.2% after a 15-Minute exposure period and 42-Hour post-exposure incubation period.
It was considered unnecessary to perform IL-1alpha analysis as the results of the MTT test were unequivocal.

Quality Criteria:
The relative mean tissue viability for the positive control treated tissues was 4.9% relative to the negative control treated tissues and the standard deviation value of the viability was 0.9%. The positive control acceptance criteria were therefore satisfied.

The mean OD562 for the negative control treated tissues was 1.293 and the standard deviation value of the viability was 3.5%. 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 11.9%. The test item acceptance criterion was therefore satisfied.

Mean OD562 Values and Viabilities for the Negative Control Item, Positive Control Item and Test Item


Item

OD562 of tissues

Mean OD562 of triplicate tissues

+/- SD of OD562

Relative individual tissue viability (%)

Relative mean viability (%)

+/- SD of Relative mean viability (%)

Negative Control Item

1.265

1.293

0.045

97.8

100*

3.5

1.345

104.0

1.269

98.1

Positive Control Item

0.076

0.063

0.011

5.9

4.9

0.9

0.058

4.5

0.055

4.3

Test Item

0.387

0.210

0.154

29.9

16.2

11.9

0.124

9.6

0.118

9.1


OD = Optical Density

SD = Standard deviation

* = The mean viability of the negative control tissues is set at 100%

Interpretation of results:
other: Skin irritant Category 2
Remarks:
according to EU CLP (EC No. 1272/2008 and its amendments).
Conclusions:
The relative mean tissue viability after 15 minutes treatment with the substance compared to the negative control tissues was 16.2%. Since the mean relative tissue viability for the substance was below 50%, the substance is considered to be a skin irritant Category 2 according to GHS.
Executive summary:

The possible skin irritation potential of the substance was evaluated using the EPISKIN reconstructed human epidermis model in compliance with OECD TG 439 and GLP principles. Triplicate skin tissues were treated by application of 10 µL undiluted test substance for an exposure period of 15 minutes. After 42 hours post-exposure incubation period, determination of the cytotoxic (irritancy) effect was performed. Cell viability was measured by enzymatic reduction of the yellow MTT tetrazolium salt (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl-tetrazolium bromide) to a blue formazan salt (within the mitochondria of viable cells) in the test substance treated tissues relative to the negative controls. The relative mean tissue viability obtained after 15 minutes treatment with the substance compared to the negative control tissues was 16.2%. Since the mean relative tissue viability for the substance was below 50% after 15 minutes treatment the substance is considered to be irritating to the skin. The positive control had a relative mean tissue viability of 4.9% after 15 minutes exposure. The standard deviation value of the percentage viability of three tissues treated identically was less than 18%, for the substance, the positive and the negative control, indicating that the test system functioned properly. Based on the result of this study the substance is considered to be irritating to the skin.

Endpoint conclusion
Endpoint conclusion:
adverse effect observed (irritating)

Eye irritation

Link to relevant study records

Referenceopen allclose all

Endpoint:
eye irritation: in vitro / ex vivo
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Study period:
2018
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Read-across information.
Justification for type of information:
The read across rationale is presented in the related Endpoint summary, the accompanying files are also attached there.
Reason / purpose for cross-reference:
read-across source
Irritation parameter:
percent corneal swelling
Run / experiment:
Slit lamp observation
Value:
0
Negative controls validity:
valid
Positive controls validity:
valid
Irritation parameter:
cornea opacity score
Run / experiment:
Slit lamp observation
Value:
0.5
Negative controls validity:
valid
Positive controls validity:
valid
Irritation parameter:
fluorescein retention score
Run / experiment:
Slit lamp observation
Value:
0
Negative controls validity:
valid
Positive controls validity:
valid
Interpretation of results:
other: Not an eye irritant
Remarks:
according to EU CLP (EC No. 1272/2008 and its amendments).
Conclusions:
The substance is not an eye irritant based on read across from Myrcenyl acetate 'mono', which was tested in an OECD TG 439.
Endpoint:
eye irritation: in vitro / ex vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
22-03-2016 to 04-04-2016
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Justification for type of information:
The information is used for read across to Pseudo linalyl acetate.
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, 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:
TEST ANIMALS
- Source: Slaughterhouse v.d. Bor, Nijkerkerveen, The Netherlands
- Age at study initiation: approximately 7 weeks
- Weight at study initiation: approximately 1.5 - 2.5 kg
-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.
- The preparation and validation of the eyes prior to the ICE-test were all according to OECD guideline 438.
Vehicle:
unchanged (no vehicle)
Controls:
other: Positive controls: Benzalkonium Chloride. Negative control: Physiological saline
Amount / concentration applied:
TEST MATERIAL
- Amount applied: 30 µL
Duration of treatment / exposure:
10 seconds
Duration of post- treatment incubation (in vitro):
240 minutes
Number of animals or in vitro replicates:
3 eyes
Details on study design:
REMOVAL OF TEST SUBSTANCE
- Washing: The eyes were rinsed with 20 mL saline
- Time after start of exposure: 10 seconds

SCORING SYSTEM: According to OECD 438 guideline. Examination of the eyes after 0, 30, 75, 120, 180, and 240 minutes

TOOL USED TO ASSESS SCORE: All examinations were carried out with the slit-lamp microscope. Fluorescein retention was only scored at approximately 30 minutes after treatment.

CONTROLS: A negative control (30 µL physiological saline) and 3 positive controls (30 µL Benzalkonium Chloride 5%) were included.
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 value
Irritation parameter:
cornea opacity score
Run / experiment:
Slit-lamp examination
Value:
0.5
Vehicle controls validity:
not applicable
Negative controls validity:
valid
Positive controls validity:
valid
Remarks on result:
other: maximum mean value
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 caused corneal effects consisting of no corneal swelling, no or slight-to-moderate opacity (mean score of 0.5) and no fluorescein retention (mean score of 0.0). 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: Microscopic examination of the corneas treated with the test substance revealed no abnormalities in two corneas. Slight erosion, moderate necrosis and very slight vacuolation of the epithelium and the epithelium partly detached from the basement membrane were observed in the third cornea, which might be related to the slight-to-moderate opacity observed in this cornea. Since observed in only one of the three cornea, these findings were considered negligible. 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 moderate or severe erosion and slight vacuolation of the epithelium, the epithelium partly detached from the basement membrane, and endothelial necrosis (two corneas).
Interpretation of results:
other: Not an eye irritant
Remarks:
according to EU CLP (EC No. 1272/2008 and its amendments).
Conclusions:
No adverse effects observed during the test. The substance is not an eye irritant.
Executive summary:

In accordance to OECD guideline 438 and GLP the test substance was examined for its in vitro eye irritating potential using the Isolated Chicken Eye (ICE) Test. In the ICE 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 caused corneal effects consisting of no corneal swelling, no or slight-to-moderate opacity (mean score of 0.5) and no fluorescein retention (mean score of 0.0). 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 revealed no abnormalities in two corneas. Slight erosion, moderate necrosis and very slight vacuolation of the epithelium and the epithelium partly detached from the basement membrane were observed in the third cornea, which might be related to the slight-to-moderate opacity observed in this cornea. Since observed in only one of the three cornea, these findings were considered negligible. 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 moderate or severe erosion and slight vacuolation of the epithelium, the epithelium partly detached from the basement membrane, and endothelial necrosis (two corneas). Based on these results, the test substance is considered to be not eye irritating.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (not irritating)

Additional information

Skin irritation and eye irritation potential is assessed based on read-across from Terpineol alpha and Myrcenyl acetate to Pseudo Linalyl Acetate. The executive summaries of the source information is presented below followed by the read-across rationale.  

Myrecenyl acetate 'mono' experimental skin irritation information

The skin irritation potential was tested using the EPISKIN reconstructed human epidermis model in compliance with OECD TG 439 and GLP principles.Triplicate skin tissues were treated by application of 10 µL undiluted test substance for an exposure period of 15 minutes.After 42 hours post-exposure incubation period, determination of the cytotoxic (irritancy) effect was performed.Cell viability was measured by enzymatic reduction of the yellow MTT tetrazolium salt (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl-tetrazolium bromide) to a blue formazan salt (within the mitochondria of viable cells) in the test substance treated tissues relative to the negative controls.The relative mean tissue viability obtained after 15 minutes treatment with the substance compared to the negative control tissues was 16.2%. Since the mean relative tissue viability for the substance was below 50% after 15 minutes treatment the substance is considered to be irritating to the skin. The positive control had a relative mean tissue viability of 4.9% after 15 minutes exposure.The standard deviation value of the percentage viability of three tissues treated identically was less than 18%, for the substance, the positive and the negative control, indicating that the test system functioned properly. Based on the result of this study the substance is considered to be irritating to the skin.

Myrcenyl acetate 'mono' experimental eye irritation information

The eye irritation was tested in an in vitro eye irritating test in accordance to OECD guideline 438 and GLP (Isolated Chicken Eye (ICE) Test). In the ICE 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 caused corneal effects consisting of no corneal swelling, no or slight-to-moderate opacity (mean score of 0.5) and no fluorescein retention (mean score of 0.0). 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 revealed no abnormalities in two corneas. Slight erosion, moderate necrosis and very slight vacuolation of the epithelium and the epithelium partly detached from the basement membrane were observed in the third cornea, which might be related to the slight-to-moderate opacity observed in this cornea. Since observed in only one of the three cornea, these findings were considered negligible. 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 moderate or severe erosion and slight vacuolation of the epithelium, the epithelium partly detached from the basement membrane, and endothelial necrosis (two corneas). Based on these results, the test substance is considered to be not eye irritating.

Skin irritation and eye irritation of Pseudo linalyl acetate using read across from Myrcenyl acetate ‘mono’ (CAS 1118-39-4)

Introduction and hypothesis for the analogue approach

Pseudo linalyl acetate has one major, two minor constituents and a number of impurities. Myrcenyl acetate ‘mono’ with 25-35% is the main constituent. Alpha and Gamma Terpinlyl acetate are the minor ones, each between 10-20%. For Pseudo linalyl acetate no skin and eye irritation information is available.In accordance with Article 13 of REACH, lacking information can be generated by means i.e. applying alternative methods such as QSARs, grouping and read-across. For assessing the skin and eye irritation potential of Pseudo linalyl acetate the analogue approach is selected because for two of its constituents, skin and eye irritation information is available which can be used for read across.

Hypothesis: Pseudo linalyl acetate has the same skin and eye irritant properties as Myrcenyl acetate ‘mono’.

Available information on skin irritation: For Myrcenyl acetate ‘mono’ an in vitro study according to OECD TG 439 and GLP (EPISKIN) is available. The relative mean tissue viability after 15 minutes treatment with the substance compared to the negative control tissues was 16.2%. Since the mean relative tissue viability for the substance was 16.2%, which is below 50%, the substance is considered to be a skin irritant Cat. 2.

Eye irritation: For Myrcenyl acetate ‘mono’ and in vitro eye irritation study is available, using the Chicken Eye test (ICE, OECDTG 438, Rel. 1). Based on the results of this test, Myrcenyl acetate ‘mono’ is considered to be not irritating to the eye.

Target chemical and source chemical(s)

Chemical structures of the target chemical and the source chemical(s) are shown in the data matrix, including physico-chemical properties and available toxicologicalinformation.

Purity / Impurities

Pseudo linalyl acetate’s key constituents are covered by Myrcenyl acetate ‘mono’. All impurities (< 10%) are not expected to impact the assessment and are presented in Appendix 1.

Analogue approach justification

According to Annex XI 1.5 read across can be used to replace testing when the similarity can be based on a common backbone and a common functional group. When using read across the result derived should be applicable for C&L and/or risk assessment and it should be presented with adequate and reliable documentation, which is presented below.

Analogue selection: For Pseudo linalyl acetate its major constituent Myrcenyl acetate ‘mono’ is selected as an analogue or which skin and eye irritation is available and which can cover the whole substance. Supporting information is available from Alpha-Terpinyl acetate.

Structural similarities and differences:All constituents of Pseudo linalyl acetate and thus including Myrcenyl acetate ‘mono’ have a hydrocarbon backbone with one or more (non-)conjugated double bonds. The hydrocarbon backbone is mostly a chain but can be cyclic as well. The constituents have an acetate ester attached to of the chain. The difference is that some constituent have a straight alkyl chain, while others are cyclic. Some impurities are alcohol derivatives of the acetate.The double conjugated bond present at the end of the alkyl chain is considered more electrophilic compared to single bonds.

Skin and eye tissue absorption: The bioavailability of Pseudo linalyl acetate and Myrcenyl acetate ‘mono’ are similar because the constituents including Myrcenyl acetate ‘mono’ have the same molecular weight (196) and a log Kow of around 4.4.

Skin and eye reactivity: For skin and eye irritation conjugated double bonds are the electrophilic fragments, which are more reactive compared to single bonds. Therefore Myrcenyl acetate ‘mono’ is a conservative representative for the skin and eye irritation. This is supported with the information from Terpinyl acetate alpha, which has only a single double bond and is not a skin or an eye irritant.

Uncertainty of the prediction: Pseudo linalyl acetate contains some alcohol impurities which may be skin or eye irritants (Alpha-Terpineol). The impurities are < 10% and therefore these do not affect the skin or eye irritation classification of Pseudo linalyl acetate.

Data matrix

The relevant information on physico-chemical properties and toxicological characteristics are presented in the Data Matrix.

Conclusions per endpoint for hazard and risk assessment

For Pseudo linalyl acetate no skin and eye irritation information is available. The available information from Myrcenyl acetate ‘mono’, being the key constituent, was selected for read across. When using read across the result derived should be applicable for C&L and/or risk assessment and be presented with adequate and reliable documentation, which has been presented above. Myrcenyl acetate ‘mono’ is a skin irritant Cat 2 but not an eye irritant, which can be directly used for read across to Pseudo linalyl acetate.

Final conclusion on hazard:Pseudo linalyl acetate is a skin irritant Cat 2 and not an eye irritant.

 

Data matrix for the read across to Pseudo linalyl acetate from Myrcenyl acetate ‘mono’ for skin and eye irritation

Common name

Pseudo linalyl acetate

Myrcenyl acetate ‘mono’

Alpha-Terpinyl acetate

Gamma-Terpinyl acetate

 

Target

Source (Major constituent)

Source supporting

(Minor constituent)

Minor constituent

Structure

Not applicable for a reaction mass

CAS #

Not applicable

1118-39-4

80-26-2

10235-63-9

EC #

944-488-1

214-262-0

201-265-7

233-564-3

REACH registration

2018

2018

Registered

Not found

Empirical formula

Not applicable

C12H20O2

C12H20O2

C12H20O2

Molecular weight

Not applicable

196.29

196

196.29

Phys-chem data

 

 

EpiSuite

EpiSuite

Log Kow

4.4 (exp.)

4.4 (exp.)

4.3

4.5

Human health endpoints

 

 

 

 

Skin irritation

 

Skin irritant Cat. 2

(Read across)

Skin irritant Cat. 2

(OECD TG 439)

Not a skin irritant

(OECD TG 404*)

Skin irritant Cat. 2

(Read across)

Eye irritation

 

Not an Eye irritant

(Read across)

Not an eye irritant

(OECDTG 438)

Not an eye irritant

(OECD TG 438*)

Not an Eye irritant

(Read across)

*Information from the ECHA website

 

Appendix 1: Constituents and possible impurities of Pseudo linalyl acetate

CAS#

Type of constituent

Name

 

 

Esters linear and alicyclic

1118-39-4

Major

2-methyl-6-methylideneoct-7-en-2-yl acetate (Myrcenyl acetate ‘mono’)

7643-61-0

Impurity

(5Z)-2,6-dimethylocta-5,7-dien-2-yl acetate

7643-62-1

Impurity

(5Z)-2,6-dimethylocta-5,7-dien-2-yl acetate

105-87-3 *

Impurity

(2E)-3,7-dimethylocta-2,6-dien-1-yl acetate (Geranyl acetate)

80-26-2 *

Minor

2-(4-methylcyclohex-3-en-1-yl)propan-2-yl acetate (Alpha-Terpinyl acetate)

150461-96-4

Impurity

1-(3,3-dimethylcyclohex-1-en-1-yl)ethyl acetate

150461-97-5

Impurity

1-(5,5-dimethylcyclohex-1-en-1-yl)ethyl acetate

10235-63-9

Minor

1-methyl-4-(propan-2-ylidene) cyclohexyl acetate (Gamma-Terpinyl acetate)

20777-47-3

Impurity

cis-1-methyl-4-(prop-1-en-2-yl)cyclohexyl acetate (Beta-terpinyl acetate)

97890-05-6

Impurity

(2Z)-2-(3,3-dimethylcyclohexylidene)ethyl acetate

76-49-3

Impurity

(1R,2S,4R)-1,7,7-trimethylbicyclo[2.2.1]hept-2-ylrel-acetate (Bornyl acetate)

210648-12-7

Impurity

3,3,5-trimethylcyclohept-4-en-1-yl acetate

 

 

Alcohol linear and alicyclic

543-39-5

Impurity

2-methyl-6-methylideneoct-7-en-2-ol (Myrcenol)

98-55-5 *

Impurity

2-(4-methylcyclohex-3-en-1-yl)propan-2-ol (Alpha-Terpineol)

586-81-2

Impurity

1-methyl-4-(propan-2-ylidene)cyclohexanol (Gamma-terpineol)

 

Justification for classification or non-classification

Based on the results, the substance needs to be classified as Skin irritation Category 2 but does not need to be classified as an eye irritant. The substance shall be labelled with 'H315: Causes skin irritation' according to EU CLP (EC No. 1272/2008 and its amendments).