Dioctyltin oxide

Endpoint summary

Description of key information

SKIN
Key study:- Warren (2012a) OECD 431, EU Method B.40, Not corrosive (in vitro, Reconstructed Human Epidermis (RHE) model)
Key study:- Warren (2012b), OECD 439, EU Method B.46, Not irritating (in vitro, Reconstructed Human Epidermis (RHE) model)
EYES
Key study:- Sanders (2012) OECD 405, EU Method B.5, Non-irritant (Rabbit, male)
Supporting study:- Warren (2012c) SkinEthic Reconstituted Human Corneal model, Non-iritant (in vitro, SkinEthic Reconstituted Human Corneal model)

Key value for chemical safety assessment

Skin irritation / corrosion

Link to relevant study records

Reference

Endpoint:

skin irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

22 August 2012 to 27 August 2012

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: The study was performed to the standardised guidelines OECD 439 and EU Method B.46 and in line GLP. The study was reported to a high standard, sufficient to assess the quality of the data presented.

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Details on test animals or test system and environmental conditions:

EPISKIN Model Kit
Supplier: SkinEthic Laboratories, Nice, France

Amount / concentration applied:

10 mg of the solid test item was applied topically.

Duration of treatment / exposure:

15 minutes

Number of animals:

Not applicable

Irritation / corrosion parameter:

other: other: OD540

Value:

0.515

Remarks on result:

other:

Remarks:

Basis: mean triplicate tissues. Time point: 15 minutes. Max. score: 0.567. Remarks: ± SD (0.066). (migrated information)

Irritation / corrosion parameter:

other: other: Relative mean viability (%)

Value:

66.7

Remarks on result:

other:

Remarks:

Basis: mean triplicate tissues. Time point: 15 minutes. Max. score: 100.0. Remarks: value in percent (%). ± SD (8.5 %). (migrated information)

Direct MTT Reduction

The MTT solution containing the test material did not turn blue which indicated that the test material did not directly reduce MTT.

Main Test - Results

The relative mean viability of the test material treated tissues was 66.7 % after a 15 minute exposure period

Table 1: Results

Item	OD540 of tissues	Mean OD540 of triplicate tissues	± SD of OD540 Relative individual tissue viability (%)	Relative individual tissue viability (%)	Relative mean viability (%)	± SD of Relaive mean viability (%)
Negative control item	0.748	0.771	0.025	97.0	100*	3.2
	0.769			99.7
	0.797			103.4
Positive control item	0.104	0.079	0.022	13.5	10.2	2.9
	0.07			9.1
	0.062			8.0
Test material	0.567	0.515	0.066	73.5	66.7	8.5
	0.441			57.2
	0.536			69.5

SD = standard deviation

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

Quality Criteria

The relative mean tissue viability for the positive control treated tissues was 10.2 % relative to the negative control treated tissues and the standard deviation value of the percentage viability was 2.9 %. The positive control criterion was therefore satisfied.

The mean OD540 for the negative control treated tissues was 0.771 and the standard deviation value of the percentage viability was 3.2 %. The negative control acceptance criterion was therefore satisfied.

The standard deviation calculated from individual percentage tissue viabilities of the three identically treated tissues was 8.5 %. The test material acceptance criterion was therefore satisfied.

Interpretation of results:

not classified

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

The test material was considered to be a 'non-irritant' to the skin.

Executive summary:

The purpose of the test was to evaluate the skin irritation potential of the test material using the EPISKIN in vitro Reconstructed Human Epidermis (RHE) Model. This method was designed to be compatible with the OECD Guideline for the Testing of Chemicals No. 439 “In Vitro Skin Irritation” (adopted 22 July 2010) and Method B.46 of Commission Regulation (EC) No. 440/2008.

 

Triplicate tissues were treated with the test material for an exposure period of 15 minutes. At the end of the exposure period the test item was rinsed from each tissue before each tissue was taken for MTT-loading. After MTT loading a total biopsy of each epidermis was made and placed into micro tubes containing acidified isopropanol for extraction of formazan crystals out of the MTT-loaded tissues. 

At the end of the formazan extraction period each tube was mixed thoroughly and duplicate 200 µL samples were transferred to the appropriate wells of a pre labelled 96 well plate. The optical density (OD) was measured at 540 nm (OD540).

Data are presented in the form of percentage viability (MTT reduction in the test item treated tissues relative to negative control tissues).

 

The relative mean viability of the test material treated tissues was 66.7 % after the 15 minute exposure period. The quality control criteria required for acceptance of results in the test were satisfied. Under the conditions of the test, the test material was concluded to be a non-irritant to the skin.

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:

5 November 2012 - 21 November 2012

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: The study was performed to standardised guidelines OECD 405 and EU Method B.5 and in line with GLP. The study was reported to a high standard, sufficient to assess the quality of the data presented.

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
- Age at study initiation: 12 - 20 weeks
- Weight at study initiation: 2.23 - 2.88 kg
- Housing: individually in suspended cages
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 17 - 23 °C
- Humidity (%): 30 - 70 %
- Air changes (per hr): at least 15 air changes per hour
- Photoperiod (hrs dark / hrs light): 12 hours dark / 12 hours light

IN-LIFE DATES: From 5 November 2012 - 21 November 2012

Vehicle:

unchanged (no vehicle)

Controls:

no

Amount / concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 0.1 mL (equivalent to approximately 78 mg)

Duration of treatment / exposure:

- Dose administration: 0.1 mL of the test material was placed into the conjunctival sac of the right eye of one rabbit, by gently pulling the lower lid away from the eyeball. The upper and lower eyelids were then held together for about 1 second immediately after treatment, to prevent loss of the test material, and then released. The left eye remained untreated and was used for control purposes. After consideration of the ocular responses in the first treated animal, a second animal was treated.

Observation period (in vivo):

Animals were observed up to 7 days post administration

Number of animals or in vitro replicates:

One male initially, followed by a further male once the irritation potential was fully assessed in the first animal.

Details on study design:

REMOVAL OF TEST SUBSTANCE
- Washing (if done): Irrigation was not performed

SCORING SYSTEM:
The reactions observed were scored in accordance with the criteria of Draize (1977).

TOOL USED TO ASSESS SCORE:
Examination of the eye was facilitated by the use of the light source from a standard ophthalmoscope.

Irritation parameter:

cornea opacity score

Basis:

animal #1

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

fully reversible

Irritation parameter:

cornea opacity score

Basis:

animal #2

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

fully reversible

Irritation parameter:

iris score

Basis:

animal #1

Time point:

24/48/72 h

Score:

0.3

Max. score:

2

Reversibility:

fully reversible within: 72 hours

Irritation parameter:

iris score

Basis:

animal #2

Time point:

24/48/72 h

Score:

0

Max. score:

2

Reversibility:

fully reversible

Irritation parameter:

conjunctivae score

Basis:

animal #1

Time point:

24/48/72 h

Score:

1.67

Max. score:

3

Reversibility:

not fully reversible within: 7 days

Irritation parameter:

conjunctivae score

Basis:

animal #2

Time point:

24/48/72 h

Score:

1

Max. score:

3

Reversibility:

fully reversible within: 7 days

Irritation parameter:

chemosis score

Basis:

animal #1

Time point:

24/48/72 h

Score:

1.3

Max. score:

4

Reversibility:

fully reversible within: 7 days

Irritation parameter:

chemosis score

Basis:

animal #2

Time point:

24/48/72 h

Score:

1

Max. score:

4

Reversibility:

fully reversible within: 7 days

Irritation parameter:

cornea opacity score

Basis:

mean

Time point:

24/48/72 h

Score:

0

Max. score:

0

Reversibility:

fully reversible

Irritation parameter:

iris score

Basis:

mean

Time point:

24/48/72 h

Score:

0.2

Max. score:

2

Reversibility:

fully reversible within: 72 hours

Irritation parameter:

conjunctivae score

Basis:

mean

Time point:

24/48/72 h

Score:

1.3

Max. score:

3

Reversibility:

fully reversible within: 7 days

Irritation parameter:

chemosis score

Basis:

mean

Time point:

24/48/72 h

Score:

1.2

Max. score:

4

Reversibility:

fully reversible within: 7 days

Irritant / corrosive response data:

No corneal effects were noted during the study. Iridial inflammation was noted in one treated eye at the 24 hour observation. Moderate conjunctival irritation was noted in both treated eyes one hour after treatment. Moderate conjunctival irritation was noted in one treated eye at 24 hours while minimal conjunctival irritation was noted in the other treated eye at 24 hours. Minimal conjunctival irritation persisted in both treated eyes to the 48 and 72 hour observations. Both treated eyes appeared normal at the 7 day observation.

Other effects:

Both animals showed expected gain in bodyweight during the study.

Table 2: Ocular Irritation Results

Animal	1					2
Time after treatment	1 h	24 h	48 h	72 h	7 d	1 h	24 h	48 h	72 h	7 d
Cornea
Degree of opacity	0	0	0	0	0	0	0	0	0	0
Area of cornea involved	0	0	0	0	0	0	0	0	0	0
Iris
Iris	0	1	0	0	0	0	0	0	0	0
Conjunctivae
Redness	2	2	2	1	0	2	1	1	1	0
Chemosis	2	2	1	1	0	2	1	1	1	0
Discharge	1	1	0	0	0	1	0	0	0	0

 

 

Interpretation of results:

not classified

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

As there was no corneal involvement in any of the two treated eyes, and mean values for the 24 to 72-Hour time points were 0.2, 1.3 and 1.2 for iritis, conjunctival redness and conjunctival chemosis, respectively, the test material is not irritating in rabbit eyes. The test material does not require classification for eye irritation in line with Regulation No. 1272/2008

Executive summary:

The eye irritation of the test material was determined in accordance with standardised guidelines OECD 405 and EU Method B.5. During the study, 0.1 mL of test material was placed into one eye of each of two rabbits and were assessed for up to 7 days to determine the grade of ocular reaction. No corneal effects were noted during the study. Iridial inflammation was noted in one treated eye at the 24 hour observation. Moderate conjunctival irritation was noted in both treated eyes one hour after treatment. Moderate conjunctival irritation was noted in one treated eye at 24 hours while minimal conjunctival irritation was noted in the other treated eye at 24 hours. Minimal conjunctival irritation persisted in both treated eyes to the 48 and 72 hour observations. All signs of irritation had completely resolved within 7 days of application.

Under the conditions of the study, the test material is not irritating in rabbit eyes. The test material does not require classification for eye irritation in line with Regulation No. 1272/2008.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Respiratory irritation

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Skin

In the key study, Warren (2012a), the corrosivity potential of the test material was determined using the validated EPISKIN in vitro Reconstructed Human Epidermis (RHE) model. The study was performed in compliance with GLP and in accordance with the standardised guidelines OECD 431 and EU Method B.40 and as such, the study was assigned a reliability score of 1 in accordance with the criteria for assessing data quality described in Klimisch (1997).

Under the conditions of the study the relative mean viability of the test item treated tissues were: 240 minutes exposure: 106.3 % ; 60 minutes exposure: 108.9 % ; 3 minutes exposure: 132.1 %. The test item was subsequently considered to be Non-Corrosive to the skin. 

 

Following on from the negative in vitro skin corrosion study, the skin irritation potential of the test material was determined in the key study Warren (2012b) using the validated EPISKIN in vitro Reconstructed Human Epidermis (RHE) model. The study was performed in compliance with GLP and in accordance with the standardised guidelines OECD 439 and EU Method B.46 and as such, the study was assigned a reliability score of 1 in accordance with the criteria for assessing data quality described in Klimisch (1997).

 Under the conditions of the study, the relative mean viability of the test material treated tissues was 66.7 % after the 15 minute exposure period. The quality control criteria required for acceptance of results in the test were satisfied. The test material was subsequently concluded to be a non-irritant to the skin.

Eye

In the supporting study, reported by Warren (2012c) the eye irritation potential of the test material was determined in a non-standardised guideline study performed in line with good scientific practices and in line with GLP. The reliability of the study was assigned a reliability score of 2 in accordance with the principles for assessing data quality in accordance with Klimisch (1997) due to the pre-validation status of the protocol. The irritation potential was assessed using the SkinEthic Reconstituted Human Corneal model (HCE, SkinEthic Laboratories, Lyon, France) after a treatment period of 10 minutes.

Solution A served as the negative control and 2 % w/v Sodium Dodecyl Sulphate served as the positive control. Under the conditions of the study the relative mean viability of the test material treated tissues after a 10 minute exposure was 102.3 %. It was, therefore, considered unnecessary to proceed with tissue histopathology. According to the protocol followed the test material was considered to be a non-irritant.

Following on from the negative in vitro eye irritation study, the eye irritation potential of the test material was determined in the key study, reported by Sanders (2012). The study was performed in line with standardised guidelines (OECD 405 and EU Method B.5) and in accordance with GLP, the study was therefore assigned in accordance with the principles for assessing data quality as described in Klimisch (1997). During the study, 0.1 mL of test material was placed into one eye of each of two rabbits and were assessed for up to 7 days to determine the grade of ocular reaction. No corneal effects were noted during the study. Iridial inflammation was noted in one treated eye at the 24 hour observation. Moderate conjunctival irritation was noted in both treated eyes one hour after treatment. Moderate conjunctival irritation was noted in one treated eye at 24 hours while minimal conjunctival irritation was noted in the other treated eye at 24 hours. Minimal conjunctival irritation persisted in both treated eyes to the 48 and 72 hour observations. All signs of irritation had completely resolved within 7 days of application. The test material does not require classification for eye irritation in line with Regulation No. 1272/2008.

Justification for selection of skin irritation / corrosion endpoint:
In vitro study to determine the skin irritation potential of the test material. Supporting information is available on skin corrosivity, however, skin irritation potential cannot be assessed on the basis of this information which is why the aforementioned study on skin irritation potential has been selected as the key study.

Justification for selection of eye irritation endpoint:
The study reported by Sanders (2010) was selected as the key study since it is an vivo study to determine eye iritation potential of test material. Supporting information is available in the form of an in-vitro study.

Justification for classification or non-classification

Skin

In accordance with with criteria for classification and labelling as defined in Regulation (EC) No 1272/2008 (CLP) and Directive 67/548/EEC (DSD), the test material does not require classification for skin irritation and corrosion.

Eye

In accordance with the criteria for classification as defined in Annex I, Regulation 1272/2008 and Directive 67/548/EEC (DSD), the test material does not require classification for eye irritation.