Registration Dossier

Administrative data

Description of key information

Skin irritation/corrosion: slightly irritating, fully reversible (rabbit)

Eye irritation/corrosion: irreversible damage (rabbit)

Key value for chemical safety assessment

Skin irritation / corrosion

Link to relevant study records
Reference
Endpoint:
skin irritation: in vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
29 january 2002 - 05 february 2002
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Remarks:
OECD TG 404
Qualifier:
according to
Guideline:
OECD Guideline 404 (Acute Dermal Irritation / Corrosion)
Version / remarks:
1992
Deviations:
no
GLP compliance:
yes
Specific details on test material used for the study:
Batch: 62630012
Expiry date: 01 November 2002
Species:
rabbit
Strain:
New Zealand White
Remarks:
Albino rabbit (SPF-Quality)
Details on test animals and environmental conditions:
Test animals
- Source: Charles River Deutschland, Kisslegg, Germany
- Age at study initiation: Animals used within the study were at least 6 weeks old
- Weight at study initiation: weights were between 1.0 and 3.5 kg
- Housing: Individually in labelled cages with perforated floors (Scanbur, Denmark, dimensions 53.5x63x38.5 cm). Acclimatisation period was at least 5 days before start of treatment under laboratory conditions.
- Diet: Standard laboratory rabbit diet (Charles River Breeding and Maintenance Diet for Rabbits,Altromin, Germany) approx. 100 g per day
- Water: Free access to tap-water

Environmental conditions
- A controlled environment was maintained in the room with optimal conditions considered as being approximately 15 air changes per hour, a temperature of 21±3°C, a relative humidity of 30-70% and 12 hours artificial fluorescent light and 12 hours dark per day.
Type of coverage:
semiocclusive
Preparation of test site:
clipped
Vehicle:
other: Ethanol
Controls:
yes, concurrent no treatment
Amount / concentration applied:
- Concentration applied: Each animal was treated by dermal application of 0.5 grams of the test substance.
- Vehicle: Since the test substance did not mix with water, the powdery test substance was moistened with watery ethanol (50% v/v), immediately before application, to ensure close contact with the animal's skin.
Duration of treatment / exposure:
4 hours
Observation period:
1, 24, 48 and 72 hours and 7 days after exposure
Number of animals:
3
Details on study design:
Treament
- Approximately 24 hours before treatment, the dorsal fur was clipped with electric clippers, exposing an area of approximately 150 square centimeters (10x15 cm^2). Whenever considered necessary, the treated skin areas were re-clipped at least 3 hours before the observations to facilitate scoring.
- A health inspection was performed prior to the commencement of treatment, to ensure that the animals were in a good state of health. Special attention was paid to the skin to be treated, which was intact and free from abnormalities.
- Each animal was treated by dermal application of 0.5 grams of the test substance. The test substance was moistened with 0.7 ml of the vehicle and applied to the skin of one flank, using a metalline patch of 2x3 cm. The patch was mounted on Micropore tape, which was wrapped around the abdomen and secured with Coban elastic bandage. Four hours after the application, the dressing was removed and the skin cleaned of residual test substance using water.

Obervation
- Mortality/Viability: twice daily
- Toxicity: at least once daily
- Body Weight: Day of treatment (prior to application)
- Irritation: The skin reactions were assessed at approximately 1, 24, 48 and 72 hours and 7 days after the removal of the dressings and test substance. The irritation scores and a description of all other (local) effects were recorded. Adjacent areas of the untreated skin of each animal served as controls

The irritation was assessed according to a numerical scoring system (according to guideline).
Irritation parameter:
erythema score
Basis:
animal: 123
Time point:
24/48/72 h
Score:
1.7
Max. score:
2
Reversibility:
fully reversible within: 7 days
Irritation parameter:
erythema score
Basis:
animal: 124
Time point:
24/48/72 h
Score:
2
Max. score:
2
Reversibility:
fully reversible within: 7 days
Irritation parameter:
erythema score
Basis:
animal: 125
Time point:
24/48/72 h
Score:
1.7
Max. score:
2
Reversibility:
fully reversible within: 7 days
Irritation parameter:
edema score
Basis:
animal: 123
Time point:
24/48/72 h
Score:
0.7
Max. score:
1
Reversibility:
fully reversible within: 7 days
Irritation parameter:
edema score
Basis:
animal: 124
Time point:
24/48/72 h
Score:
1
Max. score:
2
Reversibility:
fully reversible within: 7 days
Irritation parameter:
edema score
Basis:
animal: 125
Time point:
24/48/72 h
Score:
1
Max. score:
2
Reversibility:
fully reversible within: 7 days
Irritant / corrosive response data:
Irritation
Four hours exposure to 0.5 g of PHYTOSPHINGOSINE resulted in very slight to well-defined erythema and very slight to slight oedema in the treated skin-areas of the three rabbits. The skin irritation had resolved within 7 days after exposure in all animals. Scaliness was noted 7 days after the removal of the dressings and test substance.

Corrosion
There was no evidence of a corrosive effect on the skin.
Other effects:
Colouration
No staining of the treated skin by the test substance was observed.

Toxicity / Mortality
No symptoms of systemic toxicity were observed in the animals during the test period and no mortality occurred.

Table1. Individual skin Irritation scores.

Animal# 123 124 125
Time after exposure Erythema Oedema comments Erythema Oedema comments Erythema Oedema comments
1hour 2 1 - 2 2 - 2 2 -
24hour 2 1 - 2 1 - 2 1 -
48hour 2 1 - 2 1 - 2 1

-

72hour 1 0 - 2 1 - 1 1 -
7days 0 0 1 0 0 1 0 0 1

Table2. Mean value Irritaion scores.

Animal# Mean 24-72hrs
Erythema Oedema
123 1.7 0.7
124 2.0 1.0
125 1.7 1.0

Table3. Animal Specifications

Animal# Mean 24-72hrs
Sex Age(weeks) Body weight(grams)
123 male 7-9 1421
124 male 7-9 1651
125 male 7-9 1493
Interpretation of results:
GHS criteria not met
Remarks:
acc. to CLP Regulation
Conclusions:
The test item phytosphingosine is slightly irritating to skin. Skin irritation resolved within 7 days after exposure in all animals. There was no evidence of a corrosive effect on the skin.
Executive summary:

Primary skin irritation/corrosion of phytosphingosine was investigated in rabbit (4 -hour semiocclusive application) and carried out according to OECD 404 and in compliance with GLP.

Four hours exposure to 0.5 g of phytosphingosine resulted in very slight to well-defined erythema and very slight to slight oedema in treated skin-areas of three rabbits. Skin irritation had resolved within 7 days after exposure in all animals. Moreover, there was no evidence of a corrosive effect on the skin. In conclusion, phytosphingosine is slighlty irritating to skin.

The study was considered reliable and adequate for hazard assessment of human health.

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:
04 February 2002- 06 February 2002
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Remarks:
OECD TG 405
Qualifier:
according to
Guideline:
OECD Guideline 405 (Acute Eye Irritation / Corrosion)
Version / remarks:
1987
Deviations:
no
GLP compliance:
yes
Specific details on test material used for the study:
Batch: 62630012
Expiry date: 01 November 2002
Species:
rabbit
Strain:
New Zealand White
Remarks:
Albino Rabbit(SPF-Quality)
Details on test animals or tissues and environmental conditions:
Test animals
- Source: Charles River Deutschland, Kisslegg, Germany
- Age at study initiation: Animals used within the study were at least 6 weeks old
- Weight at study initiation: weights were between 1.0 and 3.5 kg
- Housing: Individually in labelled cages with perforated floors (Scanbur, Denmark, dimensions53.5x63x38.5 cm). Acclimatisation period was at least 5 days before start of treatment under laboratory conditions.
- Diet: Standard laboratory rabbit diet (Charles River Breeding and Maintenance Diet for Rabbits,Altromin, Germany) approx. 100 g. per day
- Water: Free access to tap-water

Environmental conditions
- A controlled environment was maintained in the room with optimal conditions considered as being approximately 15 air changes per hour, a temperature of 21±3°C, a relative humidity of 30-70% and 12 hours artificial fluorescent light and 12 hours dark per day.
Vehicle:
unchanged (no vehicle)
Amount / concentration applied:
Single samples of approximately 36 mg of PHYTOSPHINGOSINE (a volume of approximately 0.1 ml) were instilled into one eye of each of three rabbits.
Duration of treatment / exposure:
Immediately after starting the experiment
Observation period (in vivo):
Observations were made 1, 24 and 48 hours after instillation.
Duration of post- treatment incubation (in vitro):
None, based on the severity of the ocular lesions observed dunng the study, the animals were sacrificed for ethical reasons immediately after the 48 hours observation.
Number of animals or in vitro replicates:
3
Details on study design:
Treatment
A health inspection was performed prior to application of treatment to ensure that the animals were in a good state of health. Special attention was paid to the eyes, which were free from any abnormality. Each animal was treated by application of 35.8 (35.6 - 36.0) mg of the test substance (a volume of approximately 0.1 ml), in the confilactival sac of one of the eyes after gently pulling the lower lid away from the eyeball. The lids were then gently held together for about one second to prevent loss of the test substance. The other eye remained untreated and served as the reference control.
Immediately after the 24 hr observation, a solution of 2% fluorescein in water (adjusted to pH 7.0) was instilled into both yes of each animal to quantitatively determine corneal epithelial damage. Any bright green stained area, indicating epithelial damage, was estimated as a percentage of the total corneal area. To remove residual test substance, the treated eye of each animal was rinsed with approximately 50 ml tepid tap-water, using a velocity of flow which did not affect the eye, immediately after fluorescein examination on day 2. For reference control the other eye was also rinsed.

Observation
- Mortality/Viability: twice daily
- Toxicity: at least once daily
- Body Weight: Day of treatment (prior to application)
- Irritation: The eyes of each animal were examined approximately 1, 24 and 48 hours after instillation of the test substance. The irritation scores and a description of all other (local) effects were recorded.

The irritation was assessed according to the following numerical scoring system (according to guideline).

Tool used to assess score
Fluor area(%): green staining (Percentage of total corneal area) after fluorescein treatment.
Irritation parameter:
cornea opacity score
Basis:
animal #1
Time point:
24/48/72 h
Score:
2.5
Max. score:
3
Reversibility:
not reversible
Remarks on result:
probability of severe irritation
Irritation parameter:
cornea opacity score
Basis:
animal #2
Time point:
24/48/72 h
Score:
2.5
Max. score:
3
Reversibility:
not reversible
Remarks on result:
probability of severe irritation
Irritation parameter:
cornea opacity score
Basis:
animal #3
Time point:
24/48/72 h
Score:
2.5
Max. score:
3
Reversibility:
not reversible
Remarks on result:
probability of severe irritation
Irritation parameter:
iris score
Basis:
animal #1
Time point:
24/48/72 h
Score:
1
Max. score:
2
Reversibility:
not reversible
Remarks on result:
probability of severe irritation
Irritation parameter:
iris score
Basis:
animal #2
Time point:
24/48/72 h
Score:
1
Max. score:
2
Reversibility:
not reversible
Remarks on result:
probability of severe irritation
Irritation parameter:
iris score
Basis:
animal #3
Time point:
24/48/72 h
Score:
1
Max. score:
2
Reversibility:
not reversible
Remarks on result:
probability of severe irritation
Irritation parameter:
chemosis score
Basis:
animal #1
Time point:
24/48/72 h
Score:
4
Max. score:
4
Reversibility:
not reversible
Remarks on result:
probability of severe irritation
Irritation parameter:
chemosis score
Basis:
animal #2
Time point:
24/48/72 h
Score:
3.5
Max. score:
4
Reversibility:
not reversible
Remarks on result:
probability of severe irritation
Irritation parameter:
chemosis score
Basis:
animal #3
Time point:
24/48/72 h
Score:
4
Max. score:
4
Reversibility:
not reversible
Remarks on result:
probability of severe irritation
Irritation parameter:
conjunctivae score
Basis:
animal #1
Time point:
24/48/72 h
Score:
3
Max. score:
3
Reversibility:
not reversible
Remarks on result:
probability of severe irritation
Irritation parameter:
conjunctivae score
Basis:
animal #2
Time point:
24/48/72 h
Score:
3
Max. score:
3
Reversibility:
not reversible
Remarks on result:
probability of severe irritation
Irritation parameter:
conjunctivae score
Basis:
animal #3
Time point:
24/48/72 h
Score:
3
Max. score:
3
Reversibility:
not reversible
Remarks on result:
probability of severe irritation
Irritant / corrosive response data:
Irritation and Corrosion
Application of approximately 36 mg of the test substance (a volume of approximately 0.1 ml) into one eye of each of three rabbits resulted in effects on the cornea, iris and conjunctivae. Corneal injury was seen as opacity (maximum grade 3) and epithelial damage (maximum 75% of the corneal area). The corneal injury persisted until termination (at 48 hours after instillation). Iridial irritation (grade 1) was observed in all animals and persisted until termination. Irritation of the conjunctivae was seen as redness, chemosis and discharge, which persisted until termination in all animals.
At the 48 hours observation, the outside of the eyelids was red and swollen and grey/white discolouration of the eyelids and/or nictitating membrane (sign of necrosis) was noted.
Based on the severity of the ocular lesions observed during the study, the animals were sacrificed for ethical reasons immediately after the 48 hours observation.
Other effects:
Colouration
No staining of (peri) ocular tissues by the test substance was observed.
Remnants of the test substance were present in the eyes of all animals during the observation period.

Toxicity / Mortality
No symptoms of systemic toxicity were observed in the animals during the test period and no mortality occurred.

Table1: INDIVIDUAL EYE IRRITATION SCORES

Time after
dosing
Cornea Iris Conjunctivae Comments
Opacity Area Fluor area(%)   Redness Chemosis Discharge
No 116#  
1 hour 1 1 75 1 2 2 2 b
24 hours 2 3 1 3 4 3 b
48 hours 3 1 1 3 4 2 b c k 
No 118#  
1 hour 1 2 75 1 2 3 2 b
24 hours 2 3 1 3 4 2 b
48 hours 3 1 1 3 3 3 b c k l
No 119#  
1 hour 1 1 75 1 2 3 2 b
24 hours 2 3 1 3 4 2 b
48 hours 3 1 1 3 4 3 b c k l

Fluor area (%): green staining (percentage of total corneal area) after fluorescein treatment.

Comments

b: Remnants of the test substance in the eye

c: Outside of the eyelids were red and swollen

k: Grey/white discolouration of the nictitating membrane (sign of necrosis).

l: Grey/white discolouration of the eyelid (sign of necrosis)

Table2. MEAN VALUE EYE IRRITATION SCORES

Animal# Mean 24-48hrs
corneal opacity Iris Conjunctivae
Redness Chemosis
116 2.5 1.0 3.0 4.0
118 2.5 1.0 3.0 3.5
119 2.5 1.0 3.0 4.0

# Animal specifications (at start of study):

Animal no Sex Age(weeks) Body weight(grams)
116 male 10-12 2110
118 male 10-12 2335
119 male 10-12 1915
Interpretation of results:
Category 1 (irreversible effects on the eye) based on GHS criteria
Remarks:
acc. to CLP Regulation
Conclusions:
The test item phytosphingosine causes serious and irreversible damage to eye (corrosive).
Executive summary:

Acute eye irritation/corrosion of PHYTOSPHINGOSINE was investigated in the rabbit according to OECD 405 and in compliance with GLP.

Single samples of approximately 36 mg of PHYTOSPHINGOSINE (a volume of approximately 0.1 ml) were instilled into one eye of each of three rabbits. Observations were made 1, 24 and 48 hours after instillation. Application of the test substance resulted in severe adverse effects on the cornea, iris and conjunctivae. Corneal injury was seen as opacity (maximum grade 3) and epithelial damage (maximum 75% of the corneal area). The corneal injury persisted until termination (at 48 hours after instillation). Iridial irritation (grade 1) was observed in all animals and persisted until termination. Irritation of the conjunctivae was seen as redness, chemosis and discharge, which persisted until termination in all animals. Based on the severity of the ocular lesions observed during the study, the animals were sacrificed for ethical reasons immediately after the 48 hours observation. In conclusion, phytosphingosine causes serious and irreversible damage to eye (corrosive).

The study was considered reliable and adequate for hazard assessment of human health.

Based on these results and according to the classification criteria set out in the CLP Regulation, phytosphingosine is classified for Serious eye damage, Category 1, H318 "Causes serious eye damage".

Endpoint conclusion
Endpoint conclusion:
adverse effect observed (irreversible damage)

Respiratory irritation

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

Skin irritation/corrosion

Primary skin irritation/corrosion of Phytosphingosine was investigated in rabbit (4 -hour semiocclusive application) and carried out according to OECD 404 and in compliance with GLP.

Four hours exposure to 0.5 g of Phytosphingosine resulted in very slight to well-defined erythema and very slight to slight oedema in treated skin-areas of three rabbits. Skin irritation had resolved within 7 days after exposure in all animals. Moreover, there was no evidence of a corrosive effect on the skin. In conclusion, Phytosphingosine is slightly irritating to skin.

 

Eye irritation/corrosion

A reliable key study was available investigating eye irritation/corrosion of the submission substance according to OECD 405. Application of approximately 36 mg of the test substance (a volume of approximately 0.1 ml) into one eye of each of three rabbits resulted in adverse effects on the cornea, iris and conjunctivae. Based on the severity of the adverse effect observed until termination (irreversible damage). In conclusion, Phytosphingosine caused serious eye damage.

Justification for classification or non-classification

Skin irritation/corrosion

The submission substance does not need to be classified for skin irritation/corrosion according to the criteria set out in the CLP Regulation. A slight lesion was seen after exposure, but skin irritation was resolved fully after 7 days post-exposure. Therefore, Phytosphingosine does not have to be classified according to the CLP Regulation and has no obligatory labelling requirement for skin irritation.

Eye irritation/corrosion

The submission substance must be classified for eye irritation/corrosion according to the criteria set out in the CLP Regulation. Phytosphingosine is classified for Serious eye damage, Category 1, H318 "Causes serious eye damage". The classification is based on the severity of the adverse effect observed until termination (irreversible damage to the eye in all animals).