Registration Dossier

Reference

Endpoint:

skin irritation: in vivo

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Justification for type of information:

Read CSR in Section 13 for RA justification

Reason / purpose for cross-reference:

read-across source

Irritation parameter:

edema score

Basis:

animal #1

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

other: not applicable, no irritation observed

Irritation parameter:

edema score

Basis:

animal #2

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

other: no applicable. no irritation observed

Irritation parameter:

edema score

Basis:

animal #3

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

other: not applicable. no irritation observed

Irritation parameter:

primary dermal irritation index (PDII)

Basis:

animal #4

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

other: not applicable. no irritation observed

Irritation parameter:

edema score

Basis:

animal #5

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

other: not applicable. no irritation observed

Irritation parameter:

edema score

Basis:

animal #6

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

other: not applicable. no irritation observed

Irritation parameter:

erythema score

Basis:

animal #1

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

other: not applicable

Irritation parameter:

erythema score

Basis:

animal #2

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

other: not applicable

Irritation parameter:

erythema score

Basis:

animal #3

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

other: not applicable

Irritation parameter:

erythema score

Basis:

animal #4

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

other: not applicable

Irritation parameter:

erythema score

Basis:

animal #5

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

other: not applicable

Irritation parameter:

erythema score

Basis:

animal #6

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

other: not applicable

Irritant / corrosive response data:

No irritation was noted on any of the test sites during any of the scoring intervals.

Other effects:

All animals remained healthy throughout the study.

Scoring interval (h)	Overall score	Edema	Erythema
4.5	0	0	0
24	0	0	0
48	0	0	0
72	0	0	0

Interpretation of results:

not irritating

Conclusions:

Under the conditions of this study no irritation was observed in any of the animals, on any of the test sites during any of the scoring intervals. As a consequence, the test material is judged to be non-irritating to intact skin when applied topically to New Zealand White rabbits.

Executive summary:

Three male and three female New Zealand White rabbits were treated topically on each of 2 test sites which were previously clipped free of hair. Magnesium peroxide (target substance) was placed in contact with each test site. The test sites were covered with gauze patches and the trunk of each animal was wrapped with a sheet of cheesecloth. The test material was in contact with the skin for 4 h, after which the animals were unwrapped and the test material removed. Approximately 30 min after the end of the exposure period, the test sites were scored for irritation using the method of Draize. The test sites were scored daily thereafter for three days.

No irritation (edema nor erythema) was noted on any of the test sites during any of the scoring intervals.

Under the conditions of this study, the test material is judged to be non-irritating to intact skin when applied topically to New Zealand White rabbits.

As the content of magnesium peroxide in the reaction mass of magnesium carbonate and magnesium hydroxide and magnesium oxide and magnesium peroxide is lower than the content of calcium peroxide in the tested substance (37% vs. >75%, respectively), the total amount of hydrogen peroxide that can be formed is equally lower. Therefore, based on the skin irritation test result available for calcium peroxide, skin irritation effects can be excluded for the reaction mass of magnesium carbonate and magnesium hydroxide and magnesium oxide and magnesium peroxide.

Endpoint conclusion:

no adverse effect observed (not irritating)

Reference

Endpoint:

eye irritation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP study according to internationally accepted protocol. No deviations.

Qualifier:

according to guideline

Guideline:

OECD Guideline 405 (Acute Eye Irritation / Corrosion)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EPA OPP 81-4 (Acute Eye Irritation)

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: Hazleton Research Animals, Inc., Denver, Pennsylvania
- Age at study initiation: young adult
- Weight at study initiation: 2.48 - 3.82 kg
- Housing: individually, in stainless steel rabbit cages
- Diet (e.g. ad libitum): Furina High Fiber Rabbit Chow 5326, ad libitum
- Water (e.g. ad libitum): tab water, ad libitum

ENVIRONMENTAL CONDITIONS
- Temperature (°F): 65 - 74 °F
- Humidity (%): 32 - 56 %
- Photoperiod (hrs dark / hrs light): 12h fluorescent light, / 12h dark

Vehicle:

unchanged (no vehicle)

Controls:

yes, concurrent no treatment

Amount / concentration applied:

TEST MATERIAL
- Amount(s) applied: 0.10 g

Duration of treatment / exposure:

The eyes of six of he rabbits remained unwashed.
The eyes of 3 of the rabbits were gently washed with 100 mL of tap water approximately 20-30 seconds after treatment.

Observation period (in vivo):

22 days

Number of animals or in vitro replicates:

9

Details on study design:

REMOVAL OF TEST SUBSTANCE
The eyes of six of he rabbits remained unwashed. The eyes of 3 of the rabbits were gently washed with 100 mL of tap water approximately 20-30 seconds after treatment.

SCORING SYSTEM: Method of Draize

TOOL USED TO ASSESS SCORE: penlight type light, 2% sodium fluorescein

Irritation parameter:

cornea opacity score

Basis:

animal #1

Time point:

24/48/72 h

Score:

1

Max. score:

4

Reversibility:

other: 2

Irritation parameter:

cornea opacity score

Basis:

animal #2

Time point:

24/48/72 h

Score:

1

Max. score:

4

Reversibility:

other: 0

Irritation parameter:

cornea opacity score

Basis:

animal #3

Time point:

24/48/72 h

Score:

1.6

Max. score:

4

Reversibility:

other: 3

Irritation parameter:

cornea opacity score

Basis:

animal #4

Time point:

24/48/72 h

Score:

1

Max. score:

4

Reversibility:

other: 0

Irritation parameter:

cornea opacity score

Basis:

animal #5

Time point:

24/48/72 h

Score:

1

Max. score:

4

Reversibility:

other: 0

Irritation parameter:

cornea opacity score

Basis:

animal #6

Time point:

24/48/72 h

Score:

1

Max. score:

4

Reversibility:

other: 0

Irritation parameter:

iris score

Basis:

animal #1

Time point:

24/48/72 h

Score:

0.3

Max. score:

2

Reversibility:

other: 0

Irritation parameter:

iris score

Basis:

animal #2

Time point:

24/48/72 h

Score:

0.3

Max. score:

2

Reversibility:

other: 0

Irritation parameter:

overall irritation score

Remarks:

Primary Eye Irritation Index

Basis:

mean

Time point:

other: 48 h

Score:

1.3 - 28.7

Max. score:

110

Remarks on result:

other: score: washed / unwashed

Irritation parameter:

overall irritation score

Remarks:

Primary Eye Irritation Index

Basis:

mean

Time point:

other: 22 d

Score:

0 - 8.3

Max. score:

110

Remarks on result:

other: score: washed / unwashed

Irritation parameter:

overall irritation score

Remarks:

Primary Eye Irritation Index

Basis:

mean

Time point:

other: 7 d

Score:

0 - 22.8

Max. score:

110

Remarks on result:

other: score: washed / unwashed

Irritation parameter:

overall irritation score

Remarks:

Primary Eye Irritation Index

Basis:

mean

Time point:

other: 24 h

Score:

14 - 33.2

Max. score:

110

Remarks on result:

other: score: washed / unwashed

Irritation parameter:

overall irritation score

Remarks:

Primary Eye Irritation Index

Basis:

mean

Time point:

other: 1 h

Score:

10 - 23.7

Max. score:

110

Remarks on result:

other: score: washed / unwashed

Irritation parameter:

overall irritation score

Remarks:

Primary Eye Irritation Index

Basis:

mean

Time point:

other: 72 h

Score:

0 - 28.3

Max. score:

110

Remarks on result:

other: score: washed / unwashed

Irritation parameter:

iris score

Basis:

animal #3

Time point:

24/48/72 h

Score:

0

Max. score:

2

Reversibility:

other: 0

Irritation parameter:

iris score

Basis:

animal #4

Time point:

24/48/72 h

Score:

0

Max. score:

2

Reversibility:

other: 0

Irritation parameter:

iris score

Basis:

animal #5

Time point:

24/48/72 h

Score:

0

Max. score:

2

Reversibility:

other: 0

Irritation parameter:

iris score

Basis:

animal #6

Time point:

24/48/72 h

Score:

0.6

Max. score:

2

Reversibility:

other: 0

Irritation parameter:

conjunctivae score

Basis:

animal #1

Time point:

24/48/72 h

Score:

2

Max. score:

3

Reversibility:

other: 0

Irritation parameter:

conjunctivae score

Basis:

animal #2

Time point:

24/48/72 h

Score:

2

Max. score:

3

Reversibility:

other: 0

Irritation parameter:

conjunctivae score

Basis:

animal #3

Time point:

24/48/72 h

Score:

2.6

Max. score:

3

Reversibility:

other: 0

Irritation parameter:

conjunctivae score

Basis:

animal #4

Time point:

24/48/72 h

Score:

2

Max. score:

3

Reversibility:

other: 0

Irritation parameter:

conjunctivae score

Basis:

animal #5

Time point:

24/48/72 h

Score:

2.3

Max. score:

3

Reversibility:

other: 0

Irritation parameter:

conjunctivae score

Basis:

animal #6

Time point:

24/48/72 h

Score:

3

Max. score:

3

Reversibility:

other: 0

Irritation parameter:

chemosis score

Basis:

animal #1

Time point:

24/48/72 h

Score:

1.6

Max. score:

4

Reversibility:

other: 0

Irritation parameter:

chemosis score

Basis:

animal #2

Time point:

24/48/72 h

Score:

1

Max. score:

4

Reversibility:

other: 0

Irritation parameter:

chemosis score

Basis:

animal #3

Time point:

24/48/72 h

Score:

2.6

Max. score:

4

Reversibility:

other: 0

Irritation parameter:

chemosis score

Basis:

animal #4

Time point:

24/48/72 h

Score:

2.3

Max. score:

4

Reversibility:

other: 0

Irritation parameter:

chemosis score

Basis:

animal #5

Time point:

24/48/72 h

Score:

2.3

Max. score:

4

Reversibility:

other: 0

Irritation parameter:

chemosis score

Basis:

animal #6

Time point:

24/48/72 h

Score:

3

Max. score:

4

Reversibility:

other: 0

Irritant / corrosive response data:

One hour post-dosing slight corneal opacities, iritis and moderate to severe conjunctivitis was noted in unwashed eyes. Washed eyes had mild conjunctivitis. Irritation worsened after the 1-hour scoring and unwashed eyes developed pannus, hemorrhagic conjuctivae, vascularization, blanched areas on the cornea and bulging of the cornea.

Washed eyes recovered within 72 hours. However, severe irritation persisted in 2 of the unwashed eyes througout study day 22.

Cornea effects (opacity and area):

#	Wash	24h		48h		72h		mean
		Opac.	Area	Opac.	Area	Opac.	Area	Opac.	Area
1	No	1	3	1	4	1	3	1	3.3
2	No	1	2	1	2	1	2	1	2
3	No	2	4	2	2	1	4	1.6	3.3
4	No	1	2	1	2	1	2	1	2
5	No	1	1	1	3	1	2	1	2
6	No	1	3	1	3	1	3	1	3
7	Yes	1	1	0	0	0	0	0.3	0.3
8	Yes	0	0	0	0	0	0	0	0
9	Yes	1	1	0	0	0	0	0.3	0.3

Iritis:

#	Wash	24h	48h	72h	Mean
1	No	0	0	1	0.3
2	No	1	0	0	0.3
3	No	0	0	0	0
4	No	0	0	0	0
5	No	0	0	0	0
6	No	1	0	1	0.6
7	Yes	0	0	0	0
8	Yes	0	0	0	0
9	Yes	0	0	0	0

Effects on the conjuncitvae (redness, chemosis and discharge):

#	Wash	24h			48h			72h			Mean
		R	C	D	R	C	D	R	C	D	R	C	D
1	No	2	3	3	2	1	3	2	1	3	2	1.6	3
2	No	2	2	3	2	1	3	2	0	2	2	1	2.6
3	No	2	3	3	3	3	3	3	2	3	2.6	2.6	3
4	No	2	3	3	2	2	3	2	2	3	2	2.3	3
5	No	2	3	3	2	2	3	3	2	3	2.3	2.3	3
6	No	3	2	3	3	1	2	3	1	3	3	1.3	3
7	Yes	3	2	3	1	0	0	0	0	0	1	0.6	1
8	Yes	1	0	0	0	0	0	0	0	0	0.3	0	0
9	Yes	3	1	3	1	0	0	0	0	0	1.3	0.3	1

Reversibility of effects: all parameters on day 22:

#	Wash	Cornea		Iris	Conjunctivae
		Opac.	Area		R	C	D
1	No	2	2	0	0	0	0
2	No	0	0	0	0	0	0
3	No	3	2	0	0	0	0
4	No	0	0	0	0	0	0
5	No	0	0	0	0	0	0
6	No	0	0	0	0	0	0
7	Yes	0	0	0	0	0	0
8	Yes	0	0	0	0	0	0
9	Yes	0	0	0	0	0	0

Conclusions:

Under the conditions of this study , the test material is judged to be severely irritating to unwashed eyes.

Executive summary:

Calcium peroxide was instilled into the eyes of 9 New Zealand White rabbits at a dose of 0.10 g. The eyes of 6 rabbits remained unwashed while the eyes of 3 rabbits were gently washed with 100 mL of tap water approx. 20 -30 secondes after treatment. The eyes were assessed using the method of Draize and Primary Eye Irritation Indexes were calculated.

Under the conditions of this study, the test material is judged to be severely irritating to unwashed eyes.

Endpoint conclusion:

adverse effect observed (irritating)

Endpoint conclusion:

adverse effect observed (irritating)

Skin irritation

No reliable experimental data are available that assess the skin irritation effects of the reaction mass of magnesium carbonate and magnesium hydroxide and magnesium oxide and magnesium peroxide. Nevertheless, the information available for the read-across substance is considered to be sufficient for the purpose of the chemical safety assessment and for the classification and labelling.

A reliable and well-documented in vivo skin irritation study is available that assessed the skin irritation effects upon exposure of calcium peroxide on clipped rabbit skin. The study was performed according to GLP and internationally accepted protocols. No effects of irritation were observed on any of the test sites at any of the scoring intervals. As a consequence it was concluded that the test substance is not irritating to the skin.

As the content of magnesium peroxide in the reaction mass of magnesium carbonate and magnesium hydroxide and magnesium oxide and magnesium peroxide is lower than the content of calcium peroxide in the tested substance (37% vs. >75%, respectively), the total amount of hydrogen peroxide that can be formed is equally lower. Therefore, based on the skin irritation test result available for calcium peroxide, skin irritation effects can be excluded for the reaction mass of magnesium carbonate and magnesium hydroxide and magnesium oxide and magnesium peroxide.

Eye irritation

A recent in vitro eye irritation study is available on the reaction mass of magnesium carbonate and magnesium hydroxide and magnesium oxide and magnesium peroxide. It concerns a test 438 in isolated chicken’s eyes according to GLP and the OECD guideline. Three eyes were treated with 30 mg of the test item that was applied onto the center of the cornea for a duration of 10 seconds, after which the eyes were rinsed with physiological saline solution. Corneal thickness, corneal opacity and fluorescein retention were measured and any morphological effects (e.g. pitting or loosening of the epithelium) were evaluated.

The observations during the 4h observation period revealed slight corneal swelling (mean = 9.3%) and moderate cornea opacity (severity 2 on all three eyes). Slight fluorescein retention (severity 1 on two eyes and severity 2 on one eye) was noted. Test item was stuck on all cornea surfaces after the post-treatment rinse. Two cornea surfaces were cleared at 180 minutes and one cornea surfaces at 240 minutes after the post-treatment rinse. No other morphological effect was observed. Based on these observations, the isolated chicken eyes assay could not be used to conclude on the classification for effects to the eye. Therefore, histopatholoty evaluations were performed to allow concluding on the classification. One slide per each control cornea and 2 slides per each test item treated cornea were prepared. Histopathological observations were made on two sections of each of the test item treated corneas (a total of 6 sections per test item). The test item produced epithelial erosion and vacuolation in 6/6 sections. Slight erosion in 5/6 and moderate in 1/6 sections were seen. In top epithelium, very slight vacuolation in 6/6 were observed. Mid epithelium was affected with very slight vacuolation in 3/6 sections. Low epithelium was affected with very slight vacuolation in 4/6 sections. Since guideline OECD 438, 2018 for decision criteria identifying Category 1 does not describe how many slides should be examined per 1 eye, under the condition of this study with examination of 2 slides per 1 eye, this test item was classified as Category 1. However, it should be noted that the exposure duration is supposed to be 10 seconds, but the adhered test item remained for about 2 hours, so this may be an overestimation.

Based on the precautionary principle, the reaction mass of magnesium carbonate and magnesium hydroxide and magnesium oxide and magnesium peroxide is classified as Eye Damage 1.

This result is in line with the results from an in vivo eye irritation study that is available on the read-across source substance calcium peroxide. The test was performed with 9 New Zealand White rabbits. The eyes of 3 animals were washed with water approximately 20-30 seconds after treatment, whereas the eyes of the remaining 6 animals remained unwashed. The study was performed according to GLP and internationally accepted protocols. Severe effects of eye irritation were observed in the unwashed eyes, and irritation persisted beyond the 21 day observation period in two of the animals. The eyes that were washed shortly after exposure showed less severe effects, and the eye irritation subsided within 72 hours.

Respiratory irritation

No studies are available that addresses the effects of respiratory irritation following inhalation exposure to the reaction mass of magnesium carbonate and magnesium hydroxide and magnesium oxide and magnesium peroxide. However, in 1997 a study was published (Kuschner et al.) in which human volunteers were exposed to magnesium peroxide at exposure levels ranging from 5.8 to 230 mg/m³for 15 to 45 minutes. None of the subjects developed any symptoms associated with metal fume fever, nor were there any other signs of toxicity observed.

Furthermore, relevant information can be extracted from the studies available for read-across substance hydrogen peroxide in the sections on acute inhalation toxicity: a reliable acute inhalation toxicity (Hoffman, 1990) study is available for hydrogen peroxide in which male and female Sprague-Dawley rats received a single 4 -hour whole-body exposure to a 50% aqueous solution of hydrogen peroxide at the maximum attainable aerosol concentration of 170 mg/m³. All tested animals survived the exposure and subsequent 14 -day post-exposure observation period. Signs of treatment were minimal during the exposure but a few responses such as nasal discharge were noted during the whole 14 -day observation period.

Justification for selection of skin irritation / corrosion endpoint:
Reliable and well documented study on read-across substance "reaction mass of calcium carbonate and calcium dihydroxide and calcium peroxide".

Justification for selection of eye irritation endpoint:
Reliable and well documented study on the substance itself.

Effects on eye irritation: serious eye damage

Effects on respiratory irritation: irritating

In accordance to EU Classification, Labelling and Packaging of Substances and Mixtures (CLP) Regulation (EC) No. 1272/2008, classification is not necessary for skin irritation based on the available test data for read-across substance calcium peroxide in which no skin irritation effects are observed.

In accordance to EU Classification, Labelling and Packaging of Substances and Mixtures (CLP) Regulation (EC) No. 1272/2008, based on the effects observed in the available in vitro test on the reaction mass of magnesium carbonate and magnesium hydroxide and magnesium oxide and magnesium peroxide including histopathological examinations, the test substance is to be classified as Serious Eye Damage Cat. 1.

The effects of respiratory irritation observed upon exposure to hydrogen peroxide lead to a STOT-SE Cat. 3 classification for this substances. Therefore, a STOT-SE Cat. 3 with the respiratory tract as affected organ is also adopted for the reaction mass of magnesium carbonate and magnesium hydroxide and magnesium oxide and magnesium peroxide.