Registration Dossier

There is no evidence for a skin irritating potential of hexanoic acid, 3,5,5-trimethyl-, tin (2+)salt (2:1) from an in-vitro skin irritation study (OECD guideline 439) and an acute dermal toxicity study (OECD guideline 402).
Irreversible effects on the eye were observed in an eye irritation study according to OECD guideline 405, therefore hexanoic acid, 3,5,5-trimethyl-, tin (2+)salt (2:1) is classified for eye irritation category 1 / R41, according to CLP (EU-GHS) Regulation (EC) No1272/2008 and Directive 67/548/EEC, respectively.

Reference

Endpoint:

skin irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2010-11-24 to 2010-11-26

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Guideline study

Qualifier:

according to guideline

Guideline:

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

GLP compliance:

yes (incl. QA statement)

Species:

other: reconstructed human epidermis model EpiDerm™ (EPI-200-SIT)

Details on test animals or test system and environmental conditions:

EpiDerm™ is cultured from normal human keratinocytes on inert polycarbonate filters.

Amount / concentration applied:

30 µl

Duration of treatment / exposure:

Quality control 2 h (± 15 min) (Triton X-100, 1 % solution)
Positive control 60 min (± 1 min) (SDS 5 % solution)
Negative control 60 min (± 1 min) Ultra Pure Water
Test substance 60 min (± 1 min) undiluted

Observation period:

after washing process post incubation period of 42 h

Number of animals:

The test was performed in triplicate

Irritant / corrosive response data:

Test substance: mean viability 84.1 (+/- 7.2) %
Negative control: mean viability 100 (+/- 1.2) %
Positive control: mean viability 7.8 (+/- 1.3) %

Test substance: mean viability 84.1 (+/- 7.2) %

Negative control: mean viability 100 (+/- 1.2) %

Positive control: mean viability 7.8 (+/- 1.3) %

Interpretation of results:

not irritating

Remarks:

Migrated information Criteria used for interpretation of results: expert judgment

Conclusions:

Topical application of the test substance on top of the stratum corneum of the reconstructed human epidermis model EpiDerm™ (in triplicate)
resulted in a mean cell viability of 84.1 % when compared to the corresponding negative conrol. According to the evaluation scheme of OECD test
guideline 439 the test substance has to be predicted as not irritating to the skin.

Executive summary:

The skin irritating potential of the test substance hexanoic acid, 3,5,5-trimethyl-, tin (2+)salt (2:1) has been investigated using an in-vitro reconstructed human epidermal model (RHE), according to OECD guideline 439 (In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method, July 2010).

EpiDerm™ RHEs were topically exposed to 30µL of the test substance for 60 min (± 1 min). The cell viability was determined after an additional 42 h (± 2 h) post incubation period. For the analysis of the tissue viability the Standard MTT Assay was used. The viability of the negative control (UPW) was set to 100%.

The topical application of the test substance to stratum corneum of the RHE (each in triplicate) resulted in a mean cell viability of 84.1% related to the corresponding negative control.

Residual test substance adhered to the reconstructed epidermis after completion of the rinsing procedure may have had an impact on the overall test performance due to the chemically reducing properties of the test substance towards the MTT reagent. A control with freeze killed tissue, to identify possible interference, was not included in this study. However, according to the study authors the validity of the study was not affected.

The functionality of the used test system was demonstrated by topical application of Triton X-100 (1% aqueous solution) in a quality control, resulting in a mean cell viability of 95.5 % after 2 h (± 15 min) exposure. The sensitivity and accuracy of the used test system has been demonstrated with the reference substance SDS (5% aqueous solution). A mean cell viability of 7.8 % in relation to the corresponding negative control UPW was determined.

In conclusion hexanoic acid, 3,5,5-trimethyl-, tin (2+)salt (2:1) has to be predicted as not irritating to skin when the current prediction model is applied according to the international OECD Test Guideline 439.

Endpoint conclusion:

no adverse effect observed (not irritating)

Reference

Endpoint:

eye irritation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2011-04-05 to 2011-04-26

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 405 (Acute Eye Irritation / Corrosion)

Qualifier:

according to guideline

Guideline:

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

GLP compliance:

yes (incl. QA statement)

Species:

rabbit

Strain:

New Zealand White

Details on test animals or tissues and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories, Kissleg, Germany
- Age at study initiation: no data
- Weight at study initiation: 1968.0 g, 1832.4 g and 2054.8 g
- Housing: housed single in steel cages with a plastic bottom mould
- Diet (e.g. ad libitum): ad libitum Altromin 2023 Maintenance diet for rabbits
- Water (e.g. ad libitum): tap water from municipal source ad libitum
- Acclimation period: 7 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20°C ± 3°C
- Humidity (%): 30 % - 70 %
- Air changes (per hr): 8 times/hour
- Photoperiod (hrs dark / hrs light): dimmed day light

IN-LIFE DATES: From: To: 2011-03-29 to 2011-04-26

Vehicle:

unchanged (no vehicle)

Controls:

other: the left eye remained untreated and served as a control.

Amount / concentration applied:

0.1 mL of the test substance was applied by means of a syringe to the eye of the animals.

Duration of treatment / exposure:

24 hours

Observation period (in vivo):

21 days

Number of animals or in vitro replicates:

3 females

Details on study design:

REMOVAL OF TEST SUBSTANCE
- Washing (if done): The treated eyes have not been rinsed.
- Time after start of exposure: -

SCORING SYSTEM: According to OECD guideline 405, (Draize system)

TOOL USED TO ASSESS SCORE: hand-slit lamp / biomicroscope / fluorescein

After recording the observations at 24 hours, the eyes of all rabbits were further examined with the aid of fluorescein (Fluorescein sodium C.1. 45350, Merck KGaA, Darmstadt, Germany, Batch No. K 38004887 746, durable till 2013-02-28) to look for damage of the cornea. One drop of a 0.5 % (w/v) fluorescein solution in saline was poured into both eyes and washed off with saline solution 30 seconds later. Afterwards scoring was carried out by use of an UV-Iight lamp. This procedure was repeated at all observation time points until day 14.

Irritation parameter:

cornea opacity score

Basis:

animal: #1, #2, #3

Time point:

other: mean 24,48 and 72 hours

Score:

4

Max. score:

4

Reversibility:

not fully reversible within: 21 days

Irritation parameter:

iris score

Basis:

animal: #1, #2, #3

Time point:

other: mean 24,48 and 72 hours

Score:

1

Max. score:

2

Reversibility:

not fully reversible within: 21 days

Irritation parameter:

conjunctivae score

Basis:

animal: #1, #2, #3

Time point:

other: mean 24,48 and 72 hours

Score:

2

Max. score:

3

Reversibility:

not fully reversible within: 21 days

Irritation parameter:

chemosis score

Basis:

animal: #1, #3

Time point:

other: mean 24,48 and 72 hours

Score:

2.33

Max. score:

4

Reversibility:

not fully reversible within: 21 days

Irritation parameter:

chemosis score

Basis:

animal #2

Time point:

other: mean 24,48 and 72 hours

Score:

1.67

Max. score:

4

Reversibility:

not fully reversible within: 21 days

Irritant / corrosive response data:

Moderate to strong ocular reactions including iris irritation and cornea damage were observed within 24 hour following application of 0.1 ml of the
test substance to the eyes. In all animals cornea damage, iris irritation and irritating effects to the conjunctivae were irreversible within 21 days.

Other effects:

Symptoms of systemic toxicity caused by the test substance were not observed during the whole study.
Body weight gain was positive and within normal range.

Irritant/corrosive response data for each animal at each observation time

Score at time point / Reversibility	Cornea	Iris	Conjunctivae Redness	Conjunctivae Chemosis	Conjunctivae Discharge
Score at time point / Reversibility	Max. score: 4	Max. score: 2	Max. score: 3	Max. score: 4	Max.score: 3
1 h	0/0/0	1/0/1	1/2/2	1/1/1	2/2/2
24 hours	4/4/4	1/1/1	2/2/2	3/2/3	2/2/2
48 hours	4/4/4	1/1/1	2/2/2	2/2/2	2/2/2
72 hours	4/4/4	1/1/1	2/2/2	2/1/2	2/2/2
Day 7	4/4/4	1/1/1	2/2/2	1/1/2	1/1/1
Day 14	4/4/4	1/1/1	1/1/2	1/1/1	1/1/1
Day 21	2/2/2	1/1/1	1/1/1	1/1/1	0/0/0
Average 24h, 48h, 72h	4/4/4	1/1/1	2/2/2	2.33/1.67/2.33	2/2/2
Area effected (Average 24h, 48h, 72h)	4/4/4
Reversibility	not	not	not	not	not

Interpretation of results:

Category 1 (irreversible effects on the eye)

Remarks:

Migrated information Criteria used for interpretation of results: other: CLP, EU GHS (Regulation (EC) No 1272/2008)

Conclusions:

Interpretation is based on effects on the cornea, iris and conjunctiva calculated as mean scores following grading at 24, 48 and 72 hours after instillation of the test material, in consideration of reversibility. Moderate to strong effects were observed in all three animals. Cornea damage, iris irritation and slight irritation of the conjunctivae persisted until study termination (day 21 p.a.) in all animals.

Executive summary:

In a primary eye irritation study according to OECD Guideline 405 (2002) 0.1 mL of hexanoic acid, 3,5,5-trimethyl-, tin (2+)salt (2:1) was instilled into the conjunctival sac of three White New Zealand rabbits. The eyes were not rinsed after substance application. Animals then were observed for 21 days. Irritation was scored by the method of Draize. At 24, 48, 72 hours as well as 7 and 14 days post application the eyes were further examined with fluorescein to look for cornea damage.

All 3 animals showed moderate ocular reactions 1 h after application of the test substance.

24 h, 48 hand 72 h after application moderate to strong ocular reactions were observed in all three animals. In addition all rabbits showed irritation of the iris (score 1). Opaque cornea (score 4) over the whole area was observed in all animals 24 h, 48 hand 72 h after application.

Cornea damage, iris irritation and slight irritation of the conjunctivae persisted until study termination (day 21 p.a.) in all animals.

Endpoint conclusion:

adverse effect observed (irritating)

Endpoint conclusion:

no study available

Skin irritation

The skin irritating potential of the test substance hexanoic acid, 3,5,5-trimethyl-, tin (2+)salt (2:1) has been investigated using an in-vitro reconstructed human epidermal model (RHE), according to OECD guideline 439 (In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method, July 2010).

EpiDerm™ RHEs were topically exposed to 30µL of the test substance for 60 min (± 1 min). The cell viability was determined after an additional 42 h (± 2 h) post incubation period. For the analysis of the tissue viability the Standard MTT Assay was used. The viability of the negative control (UPW) was set to 100%.

The topical application of the test substance on top of stratum corneum of the RHE (each in triplicate) resulted in a mean cell viability of 84.1% related to the corresponding negative control.

Residual test substance adhered to the reconstructed epidermis after completion of the rinsing procedure may have had an impact on the overall test performance due to the chemically reducing properties of the test substance towards the MTT reagent. A control with freeze killed tissue, to identify possible interference, was not included in this study. However, according to the study authors the validity of the study was not affected.

The functionality of the used test system was demonstrated by topical application of Triton X-100 (1% aqueous solution) in a quality control, resulting in a mean cell viability of 95.5 % after 2 h (± 15 min) exposure. The sensitivity and accuracy of the used test system has been demonstrated with the reference substance SDS (5% aqueous solution). A mean cell viability of 7.8 % in relation to the corresponding negative control UPW was determined.

The test substance has to be predicted as not irritating to skin when the current prediction model is applied according to the international OECD Test Guideline 439.

Furthermore, there is no evidence for a skin irritating potential of hexanoic acid, 3,5,5-trimethyl-, tin (2+)salt (2:1) from an acute dermal toxicity study (OECD guideline 402). Groups of 5 male and female young adult white Wistar rats were dermally exposed to the test substance for 24 hours under an occlusive dressing. The applied dose of 2000 mg/kg bw is equivalent to an average concentration of 26.78 mg/cm² and 23.06 mg/cm² for males and females, respectively. Calculation is based on mean applied volume males: 0.36 mL, females: 0.31 mL; density 1.19 g/mL and area of exposure 4 x 4 cm.

No signs of irritation were recorded during the 14 day observation period.

In conclusion it can be stated that there is no evidence for a skin irritating potential of hexanoic acid, 3,5,5-trimethyl-, tin (2+)salt (2:1).

Eye irritation

Data from an in-vitro hen's egg chorioallantoic membrane test (HET-CAM), an accepted alternative method to the Draize test, to evaluate the acute irritation potential are available.

Treatment with the test substance hexanoic acid, 3,5,5-trimethyl-, tin (2+)salt (2:1) resulted in the appearance of vascular injection on the CAM of 1 egg 5 minutes post application. No other effect was detected after treatment (0.17 scores of total 21 possible scores). The irritation index of negative control tap water and positive reference substance Texapon ASV 70 Spezial 5% (w/w) were 0.0 and 10.3 of 21 total possible scores, respectively.

According to this study the test substance hexanoic acid, 3,5,5-trimethyl-, tin (2+)salt (2:1) can be graded as not irritating (0.17 scores of total 21 possible scores).

However, oppositional data from an in-vivo primary eye irritation study according to OECD Guideline OECD Guideline 405 (2002) are available.

In this study 0.1 mL of hexanoic acid, 3,5,5-trimethyl-, tin (2+)salt (2:1) was instilled into the conjunctival sac of three White New Zealand rabbits. Irritation was scored by the method of Draize during the 21 day observation period. At 24, 48, 72 hours as well as 7 and 14 days post application the eyes were further examined with fluorescein to look for cornea damage.

All 3 animals showed moderate ocular reactions 1 h after application of the test substance.

24 h, 48 hand 72 h after application moderate to strong ocular reactions were observed in all three animals. In addition all rabbits showed irritation of the iris (score 1). Opaque cornea (score 4) over the whole area was observed in all animals 24 h, 48 hand 72 h after application. Cornea damage, iris irritation and slight irritation of the conjunctivae persisted until study termination (day 21 p.a.) in all animals.

In conclusion the test substance has to be evaluated as severely irritating including irreversible effects on the eye.

Justification for selection of skin irritation / corrosion endpoint:
Data from a GLP compliant in-vitro guideline study with reliability 1 and data from an acute dermal toxicity study (GLP, RL1).

Justification for selection of eye irritation endpoint:
Data from a GLP compliant guideline study with reliability 1.

Effects on eye irritation: corrosive

For hexanoic acid, 3,5,5-trimethyl-, tin (2+)salt (2:1) a non-classification for skin irritation and Category 1 (irreversible effects on the eye) and R41 for eye irritation is justified according to CLP (EU-GHS) Regulation (EC) No1272/2008 and Directive 67/548/EEC, respectively.

There is no evidence for a skin irritating potential from an in-vitro skin irritation study according to OECD guideline 439. Furthermore, no skin irritation was observed in an acute dermal toxicity with hexanoic acid, 3,5,5-trimethyl-, tin (2+)salt (2:1).

In conclusion a non-classification with respect to skin irritation is justified.

Classification of eye irritation is based on in-vivo data from an eye irritation study according to OECD Guideline 405. Effects on the cornea, iris and conjunctiva calculated as mean scores following grading at 24, 48 and 72 hours after instillation of test substance, in consideration of reversibility were taken into account.

Moderate to strong ocular reactions were observed in all three animals, including cornea mean score of 4 in all animals. Cornea damage, iris irritation and slight irritation of the conjunctivae persisted until study termination (day 21 p.a.) in all animals.