3-methyl-1,5-pentanediyl diacrylate

Endpoint summary

Description of key information

Two in vitro studies (Valin 2013) are available to evaluate the skin irritation potential of 3-methyl-1,5-pentanediyl diacrylate : The in vitro irritation test is positive and the in vitro corrosion test is negative; Therefore 3-methyl-1,5-pentanediyl diacrylate is considered to be a skin irritating.
An in vivo eye irritation study is available on rabbits: 3-methyl-1,5-pentanediyl diacrylate is considered to be an eye irritating. 
Moreover respiratory irritation was observed in the acute toxicity by inhalation.

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:

15 April 2013 - 29 April 2013

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

other: EU Method B.46 (In vitro dermal irritation)

Deviations:

yes

Remarks:

see below

Qualifier:

according to guideline

Guideline:

other: OECD Guideline 439 (In vitro dermal irritation)

Deviations:

no

Principles of method if other than guideline:

The study was performed in accordance with study plan with the following deviations from the agreed study plan:
. § Test for direct MTT reduction with the test item: due to a typing error in the study plan, the volume of MTT solution used for the negative control in the preliminary test for direct MTT reduction which is specified in the study plan is incorrect. Indeed, the volume of MTT solution used in this study was 2 mL (as described in CiToxLAB’s SOP) instead of 2.2 mL. This deviation was considered not to have compromised the validity or integrity of the study,

GLP compliance:

yes (incl. QA statement)

Test system:

human skin model

Source species:

human

Cell type:

other: reconstructed human epidermis

Details on animal used as source of test system:

EpiskinTM Model Kit (0.38 cm2 tissues) supplied by SkinEthic Laboratories, Lyon, France.
Medium and Incubation T°C: 37°C
Dates of experimental phase: from 23 April 2013 to 29 April 2013

Vehicle:

unchanged (no vehicle)

Details on test system:

REMOVAL OF TEST SUBSTANCE
- Rinsing: at the end of the treatment period, each tissue was removed from the well of the treatment plate, and rinsed with D-PBS. Rinsing was achieved by gently filling and emptying several times each tissue with D-PBS to gently remove any residual test or control items. Excess D-PBS was removed by blotting the bottom of the tissue culture insert with absorbent paper.
The rinsed tissues were transferred to the second column of 3 wells containing 2 mL of maintenance medium in each well and the plates were incubated at +37°C, 5% CO2 in a humidified incubator for 42 (± 1) hours.

POSITIVE CONTROL
Sodium Dodecyl Sulphate (SDS) at a 5% (w/v) aqueous solution.

NEGATIVE CONTROL
Name: Dulbecco’s Phosphate-Buffered Saline (D-PBS).

SCORING SYSTEM:
- Optical density (OD) was measured at 570 nm:
Relative mean viability (%) = 100 x mean cOD(test item) / mean cOD(negative control)
where:
- mean cOD Negative Control = mean ODNC – mean ODblank
- mean cOD Test Item = mean ODTI – mean ODblank

Interpretation: see below

Control samples:

yes, concurrent no treatment

yes, concurrent positive control

Amount/concentration applied:

Amount applied per tissue: 10 ± 2 µL

Duration of treatment / exposure:

Exposure period of 15 minutes, followed by a rinsing

Duration of post-treatment incubation (if applicable):

MTT-loading after a 42h-incubation period following rinsing. Observation of MTT-> formazan transformation by viable cells, after a 3-hour MTT incubation

Number of replicates:

Triplicate for each tested substance (test item, negative control, positive control).

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

15 minute exposure (treated)

Value:

6

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Other effects / acceptance of results:

Preliminary tests
In the preliminary tests, the test item was found not to have direct MTT reducing properties or a colouring potential.

Main test
All acceptance criteria for the negative and positive controls were fulfilled. The study was therefore considered to be valid.

Following a 15 minutes exposure and a 42-hour recovery period, the relative mean viability of the tissues treated with the test item was 6% with a standard deviation of 1%. As the mean viability was < 50% after the MTT reduction, the results met the criteria for an in vitro classification as irritant (R38) to skin.

Interpretation of results:

Category 2 (irritant) based on GHS criteria

Conclusions:

Under the experimental conditions of this study, the test item is considered to be irritant to skin.

Executive summary:

The objective of this study was to evaluate the skin irritation potential of the test item using the Episkin^TMreconstructed human epidermis model.

The study design was based upon international guidelines (OECD Guideline No. 439 and Commission Regulation (EC) No. 761/2009, B.46). The study was conducted in compliance with CiToxLAB’s standard operating procedures and the principles of Good Laboratory Practice.

 

Methods

 

Preliminary tests were performed to detect the ability of the test item to directly reduce MTT as well as its colouring potential.

Following the preliminary tests, the skin irritation potential of the test item was tested in the main test. The test item and both the negative and positive controls were topically applied on triplicate tissues and incubated at room temperature for 15 (± 1) minutes. At the end of the treatment period, each tissue was rinsed with D-PBS and incubated for 42 (± 1) hours at, 5% CO₂ in a humidified incubator. The cell viability was then assessed by means of the colourimetric MTT reduction assay.

Relative viability values were calculated for each tissue and expressed as a percentage of the mean viability of the negative control tissues which was set at 100% (reference viability).

 

Results

 

Preliminary tests

In the preliminary tests, the test item was found not to have direct MTT reducing properties or a colouring potential.

Main test

All acceptance criteria for the negative and positive controls were fulfilled. The study was therefore considered to be valid.

Following a 15 minutes exposure and a 42-hour recovery period, the relative mean viability of the tissues treated with the test item was 6% with a standard deviation of 1%. As the mean viability was < 50% after the MTT reduction, the results met the criteria for an in vitro classification as irritant (R38) to skin.

 

Conclusion

 

Under the experimental conditions of this study, the test item is considered to be irritant to skin.

 

According to the results of this study, the classification of the test item should be:

- irritant (R38) (Directive 67/548/EEC) and category 2 (H315) (Regulation (EC) No. 1272/2008).

Reference 2

Endpoint:

skin corrosion: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

22 May 2013 - 13 June 2013

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

other: OECD Guideline 431 (In vitro human skin corrosion)

Deviations:

no

Qualifier:

according to guideline

Guideline:

other: EU Method B.40 bis (In vitro human skin corrosion)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test system:

human skin model

Source species:

human

Cell type:

other: reconstructed human epidermis

Details on animal used as source of test system:

Episkin TM Model Kit (0.38 cm2 tissues) supplied by SkinEthic Laboratories, Lyon, France.
Medium and Incubation T°C: 37°C
Dates of experimental phase: from 04 June 2013 to 13 June 2013

Vehicle:

unchanged (no vehicle)

Details on test system:

REMOVAL OF TEST SUBSTANCE
- Rinsing: For all treated tissues (test item-treated, positive and negative controls), at the end of the designated incubation periods, each tissue insert was removed from the well of the treatment plate, and rinsed with D-PBS.
Rinsing was achieved by gently filling and emptying several times each tissue with D-PBS to gently remove any residual dose formulation.

POSITIVE CONTROL
Name: glacial acetic acid.
The supplier and batch number of the positive control used are documented in the study files.

NEGATIVE CONTROL
Name: 0.9% NaCl.
The supplier and batch number of the negative control used are documented in the study files.

SCORING SYSTEM:
- Optical density (OD) was measured at 570 nm:
Relative mean viability (%) = 100 x mean cOD(test item) / mean cOD(negative control)
where:
- mean cOD Negative Control = mean ODNC – mean ODblank
- mean cOD Test Item = mean ODTI – mean ODblank

Interpretation: see below

Control samples:

yes, concurrent negative control

yes, concurrent positive control

Amount/concentration applied:

Amount applied per tissue: 50 µL

Duration of treatment / exposure:

For the test item, exposure periods of 3 minutes, 60 and 240 minutes, followed by rinsing.
For the positive and negative controls, exposure period of 240 minutes only, followed by rinsing.

Duration of post-treatment incubation (if applicable):

MTT-loading after rinsing. Observation of MTT-> formazan transformation by viable cells, after 3-hour MTT incubation.

Number of replicates:

Duplicate tissues for each treatment period and tested substance (test item, negative control, positive control)

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

3 minutes exposure (treated)

Value:

102

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

1 hour exposure (treated)

Value:

115

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

4 hours exposure (treated)

Value:

126

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Other effects / acceptance of results:

Preliminary test
As the MTT solution color did not turn blue/purple when compared to the negative control, the test item is presumed not to reduce MTT. As a result, no additional controls were performed on water-killed tissues in parallel to the main test.
 
Main test
All acceptance criteria for the negative and positive controls were fulfilled, therefore the study was considered to be valid.
At the end of the MTT incubation period, any tissue discoloration was evaluated with the naked eye. A blue discoloration of the test item-treated tissues following the 3 minutes, 1 hour and 4 hours exposure periods was noted. This discoloration was representative of viable tissues.
 
The relative mean viabilities of the test item-treated tissues were calculated for each time exposure treatment:
- 102% for the 3 minutes exposure,
- 115% for the 1 hour exposure,
- 126% for the 4 hours exposure.
 
These values being higher than 35%, the criteria for a non corrosive response were reached.

Interpretation of results:

other: not corrosive

Conclusions:

The test item tested in its original form, is considered to be non corrosive to the skin.

Executive summary:

The objective of this study was to evaluate the skin corrosive potential of the test item using the Episkin^TMreconstructed human epidermis model.

The study design was based upon international guidelines (OECD Guideline No. 431 and Commission Regulation (EC) No. 440/2008, B.40bis). The study was conducted in compliance with CiToxLAB’s standard operating procedures and the principles of Good Laboratory Practice.

 

Methods

 

A preliminary test was performed to detect the ability of the test item to interfere with cell viability measurements by directly reducing MTT.

 

Following the preliminary test, the corrosive potential of the test item was tested in the main assay. The test item, the negative and positive controls were topically applied on duplicate tissues and incubated at room temperature as follows: positive and negative controls for 4 hours (± 5 minutes); test item for 3 minutes (± 5 seconds), 1 hour (± 5 minutes) and 4 hours (± 5 minutes).

 

At the end of the designated incubation periods, each tissue was rinsed with D-PBS. The cell viability was then assessed by means of the MTT reduction assay.

Relative viability values were calculated for each tissue and expressed as a percentage of the mean viability of the negative control tissues which was set at 100% (reference viability).

 

Results

 

Preliminary test

As the MTT solution color did not turn blue/purple when compared to the negative control, the test item is presumed not to reduce MTT. As a result, no additional controls were performed on water-killed tissues in parallel to the main test.

 

Main test

All acceptance criteria for the negative and positive controls were fulfilled, therefore the study was considered to be valid.

At the end of the MTT incubation period, any tissue discoloration was evaluated with the naked eye. A blue discoloration of the test item-treated tissues following the 3 minutes, 1 hour and 4 hours exposure periods was noted. This discoloration was representative of viable tissues.

 

The relative mean viabilities of the test item-treated tissues were calculated for each time exposure treatment:

- 102% for the 3 minutes exposure,

- 115% for the 1 hour exposure,

- 126% for the 4 hours exposure.

 

These values being higher than 35%, the criteria for a non corrosive response were reached.

 

Conclusion

Under the experimental conditions of this study, the test item tested in its original form, is considered to be non corrosive to the skin.

 

According to these results, the classification of the test item is the following:

- non corrosive (Directive 67/548/EEC, ECE/TRANS/215 (Vol. I) and Regulation 1272/2008/EEC).

Endpoint conclusion

Endpoint conclusion:

adverse effect observed (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:

18 June 2013 -22 July 2013

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 405 (Acute Eye Irritation / Corrosion)

Deviations:

no

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Species:

rabbit

Strain:

New Zealand White

Details on test animals or tissues and environmental conditions:

TEST ANIMALS
- Source: breeder: CEGAV, Argenvilliers, France
- Age at study initiation: 2 to 4 months old at the beginning of the study
- Mean body weight at study initiation: 3730 g and 4240 g
- Fasting period before study: no
- Housing: individually housed in noryl cages
- Diet: 110 pelleted diet (free access)
- Water: tap water filtered with a 0.22 µm filter (free access)
- Acclimation period: at least 5 days before the beginning of the study.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18 ± 3°C
- Humidity (%): 50 ± 20%
- Air changes (per hr): approximately 12 cycles/hour of filtered, non-recycled air
- Photoperiod (hrs dark / hrs light): 12 h/12 h

IN-LIFE DATES: 02 July 2013 to 22 July 2013

Vehicle:

unchanged (no vehicle)

Controls:

other: untreated right eye served as a control

Amount / concentration applied:

TEST MATERIAL
- Amount(s) applied: 0.1 mL/animal

Duration of treatment / exposure:

Not applicable: single application not followed by rinsing.

Observation period (in vivo):

1, 24, 48 and 72 h after administration of the test item; if relevant, daily until reversibility of reactions

Number of animals or in vitro replicates:

two males

Details on study design:

REMOVAL OF TEST SUBSTANCE: No

SCORING SYSTEM: Draize scale.

- Conjunctival chemosis (lids and/or nictitating membranes):
0 no swelling
1 any swelling above normal (includes nictitating membranes)
2 obvious swelling with partial eversion of lids
3 swelling with lids about half-closed
4 swelling with lids more than half-closed

- Conjunctival redness (palpebral and bulbar conjunctivae, cornea and iris):
0 blood vessels normal
1 a number of blood vessels definitely hyperemic (injected)
2 diffuse, crimson colour, individual vessels not easily discernible
3 diffuse, beefy red

- Iris lesions
0 normal
1 markedly deepened rugae, congestion, swelling, moderate circum-corneal hyperemia, or injection, any of these or combination of any thereof, iris still reacting to light (sluggish reaction is positive)
2 no reaction to light, haemorrhage, gross destruction (any or all of these)

- Cornea intensity of opacity (direct examination and, if necessary, with an UV lamp)
0 no ulceration or opacity
1 scattered or diffuse areas of opacity (other than slight dulling or normal lustre), details of iris clearly visible
2 easily discernible translucent area, details of iris slightly obscured
3 nacreous areas, no details of iris visible, size of pupil barely discernible
4 opaque cornea, iris not discernible through the opacity

- Cornea area of opacity (direct examination and, if necessary, with an UV lamp)
1 one quarter (or less) but not zero
2 greater than one quarter but less than a half
3 greater than one half but less than three quarters
4 greater than three quarters up to whole area

- Any other lesions observed were noted

TOOL USED TO ASSESS SCORE: UV lamp after instillation of 0.5% sodium fluorescein solution

Irritation parameter:

chemosis score

Basis:

animal #1

Time point:

24/48/72 h

Score:

3

Max. score:

4

Reversibility:

not fully reversible within: day 21

Irritation parameter:

conjunctivae score

Remarks:

(redness)

Basis:

animal #1

Time point:

24/48/72 h

Score:

2

Max. score:

2

Reversibility:

not fully reversible within: day 12

Irritation parameter:

iris score

Basis:

animal #1

Time point:

24/48/72 h

Score:

0.7

Max. score:

1

Reversibility:

fully reversible within: day 10

Irritation parameter:

cornea opacity score

Remarks:

(intensity)

Basis:

animal #1

Time point:

24/48/72 h

Score:

1.7

Max. score:

3

Reversibility:

fully reversible within: day 7

Irritation parameter:

chemosis score

Basis:

animal #2

Time point:

24/48/72 h

Score:

3

Max. score:

3

Reversibility:

fully reversible within: day 14

Irritation parameter:

conjunctivae score

Remarks:

(redness)

Basis:

animal #2

Time point:

24/48/72 h

Score:

3

Max. score:

3

Reversibility:

fully reversible within: day 12

Irritation parameter:

iris score

Basis:

animal #2

Time point:

24/48/72 h

Score:

1

Max. score:

1

Reversibility:

fully reversible within: day 9

Irritation parameter:

cornea opacity score

Remarks:

(intensity)

Basis:

animal #2

Time point:

24/48/72 h

Score:

2

Max. score:

2

Reversibility:

fully reversible within: day 8

Irritant / corrosive response data:

First animal:
In the left treated eye, moderate chemosis of the conjunctiva was noted on days 1 and 2, becoming marked on day 3 and severe from days 4 to 6. Then, marked chemosis was observed again on days 7 to 9, becoming moderate on day 10 and slight on days 11 up to 20. Moderate redness of the conjunctiva was observed from day 1 to day 9, becoming slight until day 11.
Iris lesions was noted from day 3 and persisted up to day 9.
Moderate corneal opacity was recorded on day 3, becoming marked on day 4 up to day 6.
A moderate alopecia was observed around the eye on days 9 to 19, becoming slight on day 20.
Lacrimation was observed from days 3 to 11, associated with a white thick discharge from days 4 to 6 and neovascularisation from days 5 to 7.

Second animal:
In the left treated eye, moderate chemosis of the conjunctiva was noted on day 1, becoming marked on day 2 up to day 7 and moderate again from days 8 to 10. Then, a slight chemosis persisted up to day 13.
Moderate redness of the conjunctiva was observed on day 1, becoming marked from days 2 to 4 and moderate again from days 5 to 8. Then, a slight redness persisted up to day 11.
Iris lesions were noted on day 2 and persisted up to day 8.
Moderate corneal opacity was recorded on days 2 to 6, becoming slight on day 7. Then it completely disappeared.
A slight alopecia was observed around the eye on days 3 to 12, becoming moderate on days 13 and 14.
Lacrimation was observed on days 3 to 6, associated with a whitish thick discharge from days 4 to 6.
 
Mean scores calculated for each animal over 24, 48 and 72 hours were as follows:
- chemosis: 3.0 and 3.0;
- redness of the conjunctiva: 2.0 and 3.0;
- iris lesions: 0.7 and 1.0;
- corneal opacity: 1.7 and 2.0.
 
All these scores were indicative of eye irritation.

Other effects:

No unscheduled deaths occurred during the study and no clinical signs indicative of systemic toxicity were noted in any animals.
The body weight of the animals was unaffected by the test item treatment.

Interpretation of results:

Category 2A (irritating to eyes) based on GHS criteria

Conclusions:

Under the experimental conditions of this study, the test item was severely irritant when administered by ocular route to rabbits.

Executive summary:

The objective of this study was to evaluate the potential irritant properties of the test item for the eye following a single administration to rabbits.

This study was conducted in compliance with the principles of Good Laboratory Practice.

 

Methods

 

The test item was first administered to a single male New Zealand White rabbit.

 

As mean value from grading at 24, 48 and 72 hours after instillation was = 2 for conjunctival edema (chemosis) and for conjunctival redness, and = 1 for corneal opacity, the test item was administered in the left eye of a second animal.

 

After administration to the second animal,as mean value from grading at 24, 48 and 72 hours after instillation was = 2 for conjunctival edema (chemosis) and for conjunctival redness, and = 1 for iris lesions and for corneal opacity, no other animal was treated.

 

The test item was administered inthe conjunctival sac of the left eye. The right eye remained untreated and served as control.

A dosage-volume of 0.1 mL/animal was used.

 

For both animals, a local anesthetic was used prior to treatment.

 

The eyes were not rinsedbefore 24-hour reading.

 

Each animal was observed at least once a day for mortality and clinical signs.

Ocular reactions were observed approximately 1, 24, 48 and 72 hours after the administration and then daily until the reversibility of the ocular reactions. The mean values of the scores for chemosis, redness of the conjunctiva, iris lesions and corneal opacity were calculated for each animal. Body weight was recorded on the day of treatment and at the end of the evaluation of ocular reactions.

On completion of the observation period, the animals were sacrificed then discarded without macroscopic post-mortem examination.

Results

 

No unscheduled deaths occurred during the study and no clinical signs indicative of systemic toxicity were noted in any animals.

The body weight of the animals was unaffected by the test item treatment.

 

First animal:

In the left treated eye, moderate chemosis of the conjunctiva was noted on days 1 and 2, becoming marked on day 3 and severe from days 4 to 6. Then, marked chemosis was observed again on days 7 to 9, becoming moderate on day 10 and slight on days 11 up to 20. Moderate redness of the conjunctiva was observed from day 1 to day 9, becoming slight until day 11.

Iris lesions was noted from day 3 and persisted up to day 9.

Moderate corneal opacity was recorded on day 3, becoming marked on day 4 up to day 6.

A moderate alopecia was observed around the eye on days 9 to 19, becoming slight on day 20.

Lacrimation was observed from days 3 to 11, associated with a white thick discharge from days 4 to 6 and neovascularisation from days 5 to 7.

Second animal:

In the left treated eye, moderate chemosis of the conjunctiva was noted on day 1, becoming marked on day 2 up to day 7 and moderate again from days 8 to 10. Then, a slight chemosis persisted up to day 13.

Moderate redness of the conjunctiva was observed on day 1, becoming marked from days 2 to 4 and moderate again from days 5 to 8. Then, a slight redness persisted up to day 11.

Iris lesions were noted on day 2 and persisted up to day 8.

Moderate corneal opacity was recorded on days 2 to 6, becoming slight on day 7. Then it completely disappeared.

A slight alopecia was observed around the eye on days 3 to 12, becoming moderate on days 13 and 14.

Lacrimation was observed on days 3 to 6, associated with a whitish thick discharge from days 4 to 6.

 

Mean scores calculated for each animal over 24, 48 and 72 hours were as follows:

- chemosis: 3.0 and 3.0;

- redness of the conjunctiva: 2.0 and 3.0;

- iris lesions: 0.7 and 1.0;

- corneal opacity: 1.7 and 2.0.

 

All these scores were indicative of eye irritation.

 

Conclusion

 

Under the experimental conditions of this study, the test item was severely irritant when administered by ocular route to rabbits.

 

According to the criteria of CLP Regulation,the test item should be classified category 2 and assigned the signal word "warning" and the hazard statement "H319: causes serious eye irritation".

 

Endpoint conclusion

Endpoint conclusion:

adverse effect observed (irritating)

Respiratory irritation

Endpoint conclusion

Endpoint conclusion:

adverse effect observed (irritating)

Additional information

In vitro skin corrosion test (Valin 2013):

The objective of this study was to evaluate the skin corrosive potential of the test item using the EpiskinTM reconstructed human epidermis model (OECD 431). Following the preliminary test, the corrosive potential of the test item was tested in the main assay. The test item, the negative and positive controls were topically applied on duplicate tissues and incubated at room temperature as follows: positive and negative controls for 4 hours (± 5 minutes); test item for 3 minutes (± 5 seconds), 1 hour (± 5 minutes) and 4 hours (± 5 minutes). At the end of the designated incubation periods, each tissue was rinsed with D-PBS. The cell viability was then assessed by means of the MTT reduction.

At the end of the MTT incubation period, any tissue discoloration was evaluated with the naked eye. A blue discoloration of the test item-treated tissues following the 3 minutes, 1 hour and 4 hours exposure periods was noted. This discoloration was representative of viable tissues.

 The relative mean viabilities of the test item-treated tissues were calculated for each time exposure treatment: 102% for the 3 minutes exposure,115% for the 1 hour exposure, and, 126% for the 4 hours exposure.These values being higher than 35%, the criteria for a non corrosive response were reached.Under the experimental conditions of this study, the test item tested in its original form, is considered to be non corrosive to the skin.

In vitro skin irritation test (Valin 2013):

The objective of this study was to evaluate the skin irritation potential of the test item using the EpiskinTM reconstructed human epidermis model (OECD 439). Following the preliminary tests, the skin irritation potential of the test item was tested in the main test. The test item and both the negative and positive controls were topically applied on triplicate tissues and incubated at room temperature for 15 (± 1) minutes. At the end of the treatment period, each tissue was rinsed with D-PBS and incubated for 42 (± 1) hours at, 5% CO2 in a humidified incubator. The cell viability was then assessed by means of the colourimetric MTT reduction assay. Relative viability values were calculated for each tissue and expressed as a percentage of the mean viability of the negative control tissues which was set at 100% (reference viability). Following a 15 minutes exposure and a 42-hour recovery period, the relative mean viability of the tissues treated with the test item was 6% with a standard deviation of 1%. As the mean viability was < 50% after the MTT reduction, the results met the criteria for anin vitroclassification as irritant to skin.Under the experimental conditions of this study, the test item is considered to be irritating to skin.

In vivo eye irritation test (Leclere 2014):

The objective of this study was to evaluate the potential irritant properties of the test item for the eye following a single administration to rabbits (OECD 405).The test item was administered inthe conjunctival sac of the left eye. The right eye remained untreated and served as control. A dosage-volume of 0.1 mL/animal was used. For both animals, a local anesthetic was used prior to treatment. The eyes were not rinsed before 24-hour reading.

No unscheduled deaths occurred during the study and no clinical signs indicative of systemic toxicity were noted in any animals.

The body weight of the animals was unaffected by the test item treatment.

First animal:In the left treated eye, moderate chemosis of the conjunctiva was noted on days 1 and 2, becoming marked on day 3 and severe from days 4 to 6. Then, marked chemosis was observed again on days 7 to 9, becoming moderate on day 10 and slight on days 11 up to 20. Moderate redness of the conjunctiva was observed from day 1 to day 9, becoming slight until day 11.Iris lesions was noted from day 3 and persisted up to day 9. Moderate corneal opacity was recorded on day 3, becoming marked on day 4 up to day 6. A moderate alopecia was observed around the eye on days 9 to 19, becoming slight on day 20. Lacrimation was observed from days 3 to 11, associated with a white thick discharge from days 4 to 6 and neovascularisation from days 5 to 7.

Second animal:In the left treated eye, moderate chemosis of the conjunctiva was noted on day 1, becoming marked on day 2 up to day 7 and moderate again from days 8 to 10. Then, a slight chemosis persisted up to day 13. Moderate redness of the conjunctiva was observed on day 1, becoming marked from days 2 to 4 and moderate again from days 5 to 8. Then, a slight redness persisted up to day 11. Iris lesions were noted on day 2 and persisted up to day 8. Moderate corneal opacity was recorded on days 2 to 6, becoming slight on day 7. Then it completely disappeared.

A slight alopecia was observed around the eye on days 3 to 12, becoming moderate on days 13 and 14. Lacrimation was observed on days 3 to 6, associated with a whitish thick discharge from days 4 to 6.

Mean scores calculated for each animal over 24, 48 and 72 hours were as follows:

- chemosis: 3.0 and 3.0;

- redness of the conjunctiva: 2.0 and 3.0;

-iris lesions: 0.7 and 1.0;

-corneal opacity: 1.7 and 2.0.

All these scores were indicative of eye irritation.

Under the experimental conditions of this study, the test item was severely irritant when administered by ocular route to rabbits.

Respiratory irritation (acute inhalation study, TNO 2016) :

In the acute study, respiratory irritation was evident in animals exposed to 3 -methyl-1.5 -pentanediyl diacrylate. Indeed, hemarrhagic nasal discharge was observed until day 6 in five out of 6 animals exposed to 1.05 g/m3, and nasal discharge was observed on the day of exposure in 5 out of 6 animals exposed to 5.14 g/m3.

 

 

Justification for classification or non-classification

Based on the in vitro and in vivo studies, 3-methyl-1,5-pentanediyl diacrylate is irritating for skin, eyes and respiratory tract according to the Regulation EC n°1272/2008 : Skin Irrit. 2 (H315: Causes skin irritation), Eye Irrit. 2 (H319: Causes serious eye irritation), STOT SE 3 (H335: May cause respiratory irritation).