Alkenes, C11-14, hydroformylation products, distn. residues, reaction products with maleic anhydride and sodium bisulfite, sodium salts

Endpoint summary

Description of key information

An in-vitro skin irritation study, conducted according to a fully validated and accepted method OECD TG 439, has demonstrated that Alkenes hydroformylated, sulfosuccinates, sodium salt is not irritating.
An in-vitro eye irritation study on Human Corneal Epithelium (HCE) showed that Alkenes hydroformylated, sulfosuccinates, sodium salt is an eye irritant..

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

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: According to: OECD Guideline for the Testing of Chemicals 439: In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method (Original Guideline adopted July 22, 2010)

Qualifier:

according to guideline

Guideline:

other: OECD Guideline for the Testing of Chemicals 439: In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method (Original Guideline adopted July 22, 2010)

GLP compliance:

yes (incl. QA statement)

Amount / concentration applied:

Each approximately 10 mg of the neat test item were applied to three EPISKIN (Skinethic) tissues. Addtionally, the tissues were wetted with 15 µL of deionised water.

Duration of treatment / exposure:

The skin equivalents were exposed to the test item for 15 minutes. After completion of the treatment the test item was rinsed off and the skin equivalents were incuated for further 42 hours.

Details on study design:

Each approximately 10 mg of the neat test item and 10 µl of the negative control (deionised water) and the positive control (5% SLS) were applied to each three EPISKIN (Skinethic) tissues. Additionally, the test item treated skin equivalents were wetted with 15 µL of deionised water. The test item as well as the controls were rinsed off after 15 minutes treatment. After further 42 hours incubation the tissues were treated with the MTT solution for 3 hours following approximately 72 hours extraction of the colorant from the cells. The amount of extracted colorant was determined photometrically at a wavelength of 570 nm.

Irritation / corrosion parameter:

% tissue viability

Value:

93

Irritant / corrosive response data:

After treatment with the test item Alkenes hydroformylated, sulfosuccinates, sodium salt the mean relative absorbance value was reduced to 93.0%. This value is well above the threshold for irritancy of ≤ 50%. Therefore, the test item is not considered to possess an irritant potential.

Results after treatment with Alkenes hydroformylated, sulfosuccinates, sodium salt and controls

 

Dose group	Treatment Interval	Absorbance 570 nm Tissue 1*	Absorbance 570 nm Tissue 2*	Absorbance 570 nm Tissue 3*	Mean Absorbance of 3 Tissues	Relative Absorbance [%] Tissue 1, 2 + 3**	Standard Deviation [%]	Rel. Absorbance [% of Negative Control]***
Negative Control	15 min	1.106	0.909	1.011	1.009	109.6 90.1 100.2	9.8	100.0
Positive Control	15 min	0.242	0.297	0.193	0.244	24.0 29.5 19.1	5.2	24.2
Test Item	15 min	0.922	0.946	0.947	0.938	91.4 93.8 93.9	1.4	93.0

*       Mean of two replicate wells after blank correction

**      relative absorbance per tissue [rounded values]:
***    relative absorbance per treatment group [rounded values]:

Optical evaluation of the MTT-reducing capacity of the test item after 1 hour incubation with MTT-reagent did not show blue colour.

The mean relative absorbance value of the test item, corresponding to the cell viability, decreased to 93.0% (threshold for irritancy:≤50%), consequently the test item was notirritant to skin.

Interpretation of results:

not irritating

Remarks:

Migrated information Criteria used for interpretation of results: OECD GHS

Conclusions:

In conclusion, it can be stated that in this study and under the experimental conditions reported, the test item Alkenes hydroformylated, sulfosuccinates, sodium salt is not irritant to skin.

Executive summary:

This in vitro study was performed to assess the irritation potential of Alkenes hydroformylated, sulfosuccinates, sodium salt by means of the Human Skin Model Test.

Three tissues of the human skin model EpiSkin™ were treated with the test item, the negative or the positive control for 15 minutes.

Prior to the test the test item was thawed at 37 ± 1 °C and homogenised. Since the test item was very pasty afterwards, it was applied with a measuring spoon. Approximately 10 mg of the test item were applied to each tissue, wetted with 15 µL of deionised water, and spread.

The test item and the positive and negative controls were washed off the skin tissuesafter 15 minutes treatment. After further incubation for about 42 hours the tissues were treated with the MTT solution for 3 hours following approximately 72 hours extraction of the colorant from the cells. The amount of extracted colorant was determinedphotometrically at 570 nm.

10 µL of either the negative control (deionised water) or the positive control (5% Sodium lauryl sulfate) were applied to each tissue.

After treatment with the negative control the absorbance values were well within the required acceptability criterion of mean OD of greater or equal than 0.6 till less or equval 1.5 for the15 minutes treatment interval thus showing the quality of the tissues.

Treatment with the positive control induced a decrease in the relative absorbance as compared to the negative control to 24.2% thus ensuring the validity of the test system.

The standard deviations between the % variabilities of the test item, the positive and negative controls were below 10% (threshold of the "OECD TG 439 Guideline for the Testing of Chemicals 439: In vitro Skin Irritation: Reconstructed Human Epidermis Test Method”: 18%), thus ensuring the validity of the study.

After treatment with the test item Alkenes hydroformylated, sulfosuccinates, sodium salt the mean relative absorbance value was reduced to 93.0%. This value is well above the threshold for irritancy of≤50%. Therefore, the test item is not considered to possess an irritant potential.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Eye irritation

Link to relevant study records

Reference

Endpoint:

eye irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

from 20th November 2012 to 22nd November 2012

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Well documented study conducted according to a recognized method.

Qualifier:

according to guideline

Guideline:

other: SOP M04

Deviations:

not specified

Principles of method if other than guideline:

The method allows to evaluate a toxic event and related cyotoxicity by a colorimetrtc assay. Solution of MTT in balanced saline is yeliowish in color. Mitocondrial dehydrogenase of viable cells cleaves the tetrazolium ring yielding blue/purple MTT crystals which are insoluble in aqueous solutions. Crystals formed by viable cells are retained in the polycarbonate filter used as substrate for the 3D construct. Intense purple colour of the cultures indicates the viability of the cell, whereas colour remains white when necrosis occurs. Negative controls are of a dark blue colour and positive controls are white-yellow.
MTT crystals are extracted by Isopropanol and optical density Is measured at 570 nm using the TECAN INFINITE M-200 spectrophotometer. Results are expressed as percentage of cell viability compared to the negative control.

GLP compliance:

no

Species:

other: not applicable

Strain:

other: not applicable

Details on test animals or tissues and environmental conditions:

the SkinEthic Reconstituted Human Corneal Epithelium of 0.5 cm2 is used for the evaluation Epithelial human immortalized cells (HICEC) are deposed on a polycarbonate filter and cultured at the air-liquid interface for 5 days in a chemically defined medium In order to form a structured epithelium.

Vehicle:

unchanged (no vehicle)

Controls:

not required

Amount / concentration applied:

30 µl

Duration of treatment / exposure:

Eye Irritation potential of the test item was assessed at 1 h followed by product washing and a post Incubation period of 16h.

Number of animals or in vitro replicates:

not applicable

Details on study design:

Immediately after the arrival in the laboratory the HCE was removed from the agarose nutrient solution under a sterile air flow cabin The inserts were rapidly placed in a 12- well plate previously filled with 0.75 mL of the SkinEthic maintenance medium at room temperature. The wells were placed in an incubator at 37°C 5% CO2 and saturated humidity overnight. The test was started the day after the arrival.

The test was performed on triplicate tissues
The positive control was Ethanol (99,99%) and the negative control was saline solution (NaCI 0,9%) 30 µl of controls (liquids) were directly and uniformly applied topically
Alkenes, C11-l4 hydroformylation products, distn. residues reaction products with maleic anhydride and sodium bisulfite sodium salts (C11— 14) is a liquid and it was tested at the dose defined for liquid (30 µl) according to SOP M 04
Eye Irritation potential of the test item was assessed at lh followed by product washing and a post Incubation period of 16h.

Irritation parameter:

other: cell viability

Basis:

mean

Time point:

other: 1h + 16h exposure

Reversibility:

other: not applicable

Remarks on result:

other: mean tissue viability is < 50%.

Irritant / corrosive response data:

The prediction model for eye irritation classification is based on cell viability (MTT - modified protocol) after 1h + 16h exposure.
A cut-off of 50% is accepted for dicriminating between potential irritant (≤ 50%) and not classified (> 50%). Positive control (ethanol) satisies acceptance criteria if it is classified as irritant (≤ 50%).

Interpretation of results:

irritating

Remarks:

Migrated information Criteria used for interpretation of results: expert judgment

Conclusions:

According to the adopted prediction model, based on MTT results, the test item has been classified as reported irritant for the eye.

Executive summary:

An in-vitro on Human Corneal Epithelium model (HCE) was applied in order to assess the eye irritation potential of Alkenes, C11-14, hydroformylation products, distn. residues, reaction products with maleic anhydride and sodium bisulfite, sodium salts.

The HCE, Human Corneal Epithalium model, is currently under a cosmetic Europe (CosEU, x COLIPA)-ECVAM validation study (2009 -2012) as test method replacement alternative for regulatory classification (irritant/non irritant for the eye) by using the MTT test after exposure of 1h+16h of post incubation. According to the adopted prediction model, based on MTT results, the test item has been classified as reported irritant for the eye.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed (irritating)

Respiratory irritation

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Skin irritation:

A study conducted according to OECD TG 439 is available. The method is fully validated and accepted for the assessment of skin irritation potential. Negative results from this test can be used as key study to justify a non classification for skin irritation only. Alkenes hydroformylated, sulfosuccinates, sodium salt was tested with an evident negative result.

Eye irritation:

A Bovine Corneal Opacity and Permeability Test Method (BCOP) for Identifying Ocular Corrosives and severe Irritants ( OECD TG 437 ) is available. The BCOP test on Alkenes hydroformylated, sulfosuccinates, sodium salt was clearly negative as it did not cause an increase of the corneal opacity as well as permeability. This result is valid for a non-classification as severe eye irritant, but unfortunately not enough to conclude whether this test item is irritant or not.

Alkenes hydroformylated, sulfosuccinates, sodium salt was then tested on HCE with a contact time of 1h and 16hs of recovery. In such conditions, the result in term of cell viability was 0.58%, very similar to the positive control, which is ethanol and returned the value of 0.65%. According to the prediction model of this assay, this result leads to a classification as irritant. The advantage of the HCE method is that testing is performed directly on a model of human epithelium and therefore its relevance should be very high.

Additional information are reported in the field "Attached document":

Justification for selection of skin irritation / corrosion endpoint:
Test conducted according to OECD TG 439, the method is fully validated and accepted for the assessment of skin irritation potential

Justification for selection of eye irritation endpoint:
Well documented GLP study conducted according to a recognized method.

Effects on eye irritation: irritating

Justification for classification or non-classification

According to the proposed testing strategy, Alkenes hydroformylated, sulfosuccinates, sodium salt is classified as irritating to eyes (Category 2), H319: Causes serious eye irritation.

In fact, it was tested with a validated method that can exclude the classification for serious damage to the eyes (H318). The irritant activity is confirmed with HCE model, whose suitability is well demonstrated, even though formal validation is still not available.