Reaction products of tetraisopropyl titanate (4+), hydroxyalkaloic acid and substituted alkanol

Endpoint summary

Description of key information

non skin irritant

eye irritant

Key value for chemical safety assessment

Skin irritation / corrosion

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Eye irritation

Endpoint conclusion

Endpoint conclusion:

adverse effect observed (irritating)

Respiratory irritation

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Skin corrosion

The potential of the test item to be corrosive to the skin was investigated through an in vitro skin corrosion study, using a commercial reconstructed human epidermis (RhE) model named EPISKIN™. The experimental procedures are based on the OECD Guideline for testing of chemicals no. 431. The test item, as well as controls, were tested for their ability to impair cell viability after an exposure period of 3, 60 and 240 minutes. The final endpoint of the assay is the colorimetric measurement of MTT reduction (blue formazan salt) in the test system, being this reaction an index of cell viability.

Before the Main Assay, a preliminary test was carried out to evaluate the compatibility of the test item with the test system. In particular, the test item was assayed for the ability of reducing MTT and colouring water per se. No interaction was recorded between the test

item and MTT in test conditions similar to those of the Main Assay. Moreover, no colouring potential of the test item in contact with water was recorded, with an OD value of 0.048. Based on these results, no additional controls were added in the main phase.

In theMain Assay, for each treatment time, being the test item a sticky liquid, it was applied as supplied in two replicates at the treatment level of 50 ± 2 mg/epidermis unit, each measuring 0.38 cm2 (treatment level: 131.6 mg/cm2). Positive and negative controls (Glacial acetic acid and Physiological saline, respectively) were concurrently tested, in the same number of replicates and test conditions at the treatment level of 50 μL/epidermis unit. Positive control was included only at the longest treatment time of 240 minutes, while a negative control was

included for each treatment time. In theMain Assay, the negative controls gave the expected baseline value (Optical Density values ≥ 0.6 and ≤ 1.5) and variability (difference of viability between the two replicates lower than 30%), with the only exception of the negative control OD value, after 60 minutes of treatment, which slightly exceeded the maximum acceptable value. However, the results obtained were considered reliable for the overall evaluation. For each treatment time, the concurrent negative control mean value is considered the baseline value of the treatment series and thus represents 100% of cell viability. The positive control caused the expected cell death (1% of cell viability, when compared to the negative control). Based on the stated criteria, the assay was regarded as valid.

The test item did not induce cell death in any replicate, at any treatment time. Each mean cell viability, after the concurrent blank subtraction, was as follows:

Treatment time (minutes) Mean cell viability (%)

3 110

60 ± 5 104

240 ± 5 90

At 3 and 60 minute treatments, intra-replicate variability was acceptable, with a difference of viability between the two replicates lower than 30%. After 240 minutes of treatment, intrareplicate variability for test item treated tissues was slightly higher than the acceptable one;

however both mean values were largely higher than the stated threshold for corrosion activity. Therefore results were considered reliable for evaluation of corrosion potential of the test item.

Based on the results obtained, the test item is classified as non-corrosive to the skin.

Skin irritation

The in vitro study was performed in order to evaluate the potential of the test item to evoke skin irritation in a reconstructed human epidermis (RhE) test method. The test method was performed according to OECD 439 and EU-Method B.46.

Three tissues of the human skin model EpiDermTM were treated with the test item for 60 minutes. The test item was applied directly to each tissue and spread to match the tissue size (0.63 cm2). DPBS-buffer was used as negative control and 5% SDS solution was used as positive control. After exposure of the tissues with the test item, the test item was rinsed from the tissues and cell viability of the tissues were evaluated by addition of MTT, which can be reduced to formazan. The formazan production was evaluated by measuring th eoptical density (OD) of the resulting solution. All validity criteria were met. The mean value of relative tissue viability was reduced to 94.3 %. This value is above the threshold for skin irritation potential (50%). Test items that induce values above the threshold of 50% are considered non-irritant to skin. Therefore, the test item is considered non- irritant to skin in the Reconstructed human Epidermis (RhE) Test Method.

Eye irritation

The Eye Hazard Potential of the test item was evaluated in a RhCE model in an in vitro study according to OECD Guideline 492. The test item was applied to a three-dimensional human cornea tissue model in duplicate for an exposure time of 6 hours. The solid test item was applied to two tissue replicates.

After treatment, the respective substance was rinsed from the tissue; then, cell viability of the tissues was evaluated by addition of MTT, which can be reduced to formazan. The formazan production was evaluated by measuring the optical density (OD) of the resulting solution.

Demineralised water was used as negative control and methyl acetate was used as positive control.

All validity criteria were met. After treatment with the test item, the mean value of relative tissue viability was 1.8%. This value is below the threshold for eye irritation potential (≤ 60%). Under the conditions of the test, the test item is considered either eye irritant or inducing serious eye damage in the EpiOcularTM Eye Irritation Test.

Based on the in vitro results and according to the OECD Guideline 492, the EpiOcularTM Eye Irritation Test does not allow discrimination between eye irritation/reversible effects on the eye (Category 2) and serious eye damage/irreversible effects on the eye (Category 1). For these purposes, further testing with other suitable test methods is required.

The evaluation of the test system in the Bovine Corneal Opacity and Permeability (BCOP) Test Method following OECD Guideline 437 and EU Method B.47. The test item was applied onto the cornea of a bovine eye (three replicates) which had been previously incubated with cMEM without phenol red at 32 ± 1 °C for 1 hour and whose opacity had been measured. The negative control (Hank's Balanced Salt Solution (HBSS)) showed no irritating effect on the cornea and the calculated mean IVIS (In Vitro Irritancy Score) was 0.47. The positive control (20% imidazole solution) induced serious eye damage of the cornea and was within two standard deviations of the current historical mean. The calculated mean IVIS was 104.05. The test item was tested as 20% solution in HBSS and incubated on the cornea for 4 hours at 32 ± 1 °C. After removal of the test item, opacity and permeability values were measured. Under the conditions of this study, the test item showed effects on the cornea of the bovine eye. The calculated mean IVIS was 15.48.

According to OECD Guideline no. 437 (Oct. 2017), a substance with an IVIS > 3 and ≤ 55 induces effects on the cornea, that cannot be classified in an UN GHS Category for eye damage with the BCOP study only. In this case no prediction can be made.

The eye irritation of the substance is evaluated ina weight of evidence approach. Considering the results of both in vitro studies, it can be ccocncluded that the substance is an Eye irritant.

Justification for classification or non-classification

Skin irritation

Based on the in vitro results and on the OECD 439 criteria, the test item is not classified for skin irritation as per the CLP Regulation (EC) No. 1272/2008.

Eye Hazard Potential

Considering the results obtained in the OECD 492, the substance can be considered either as a substance causing eye irritation/reversible effects on the eye (Category 2) or serious eye damagae/reversible effects on th eeye (Category 1). However, considering the results obtained in the OECD 437, the classification in Cat 1 is excluded since IVIS > 3. In a weight of evidence approach, the substance is classified as Eye irritant (H319) as per the CLP Regulation (EC) No. 1272/2008.