Reaction product with n-butanol of high-boiling stream resulting from the hydroxylation, oxidative cleavage and hydrolysis of vegetable oil saturated and unsaturated C16-C22 triglycerides.

Endpoint summary

Description of key information

The substance does not cause skin irritation/corrosion nor eye irritation or dammage.

Key value for chemical safety assessment

Skin irritation / corrosion

Link to relevant study records

Reference

Endpoint:

skin irritation / corrosion

Remarks:

in vitro

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2013

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Recent GLP compliant study with detailed test report

Qualifier:

according to guideline

Guideline:

OECD Guideline 431 (In Vitro Skin Corrosion: Human Skin Model Test)

GLP compliance:

yes (incl. QA statement)

Test system:

human skin model

Source species:

other: skin model EpiDerm

Cell type:

other: skin model EpiDerm

Cell source:

other: skin model EpiDerm

Source strain:

other: skin model EpiDerm

Control samples:

yes, concurrent negative control

yes, concurrent positive control

Species:

other: No test animals were used, as it concerns an in vitro test

Strain:

other: not applicable (in vitro test)

Vehicle:

unchanged (no vehicle)

Controls:

other: Deionised water was used as negative control, KOH was used as a positive control

Amount / concentration applied:

The liquid test item was applied without preparation (50 microL to each well)

Duration of treatment / exposure:

Three minutes and one hour, respectively

Observation period:

The tissues were incubated with MTT reagent for three hours. After this time, the MTT reagent was aspirated and replaced by DPBS buffer. This was then aspirated, too, and replaced several times. At last, each insert was thoroughly dried and set into an empty, pre-warmed plate. Into each well, 2 mL isopropanole were pipetted. The plate was then covered with Parafilm and left to stand overnight at room temperature.

Number of animals:

Not applicable (in vitro test)

Details on study design:

After incubation overnight, the inserts in which formazan had been produced over night were pierced with an injection needle, taking care that all colour was extracted. The inserts were then discarded and the content of each well was thoroughly mixed in order to achieve homogenenisation. From each well, three replicates with 200 microL Solution (each) were pipetted into a 96-well plate which was read in a plate spectral photometer at 570 nm.

The photometric absorption of the negative controls was considered as 100%. For the mean of the three replicates of test item and positive control, formazan production was calculated as % photometric absorption compared with negative control.

Irritation / corrosion parameter:

other: absorption

Run / experiment:

Formazan production

Value:

ca. 1.727

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

no indication of irritation

The absorption values of negative control, test item and positive control are given in the following table:

Absorption values

 

Negative Control		Test item		Positive control		Incubation
Tissue 1	Tissue 2	Tissue 1	Tissue 2	Tissue 1	Tissue 2
1,814	2,046	1,824	1,665	0,587	0,636	3 min
1,778	1,98	1,784	1,664	0,559	0,683
1,768	1,953	1,764	1,66	0,598	0,57
2,067	1,724	1,656	1,685	0,158	0,152	1 hour
1,994	1,962	1,758	1,825	0,172	0,172
2,085	2,065	1,795	1,837	0,185	0,174
Mean		Mean		Mean
1,89		1,727		0,606		3 min
1,983		1,759		0,169		1 hour

For the test item and the positive control, the following percentage values of mean formazan production were calculated in comparison to the mean of the negative controls.

Test item	Positive control	Incubation
91,40%	32,00%	3 min
88,70%	8,50%	1 hour

 

Interpretation of results:

other: not corrosive

Remarks:

Criteria used for interpretation of results: EU

Executive summary:

The skin corrosion potential of the substance was assessed via an in vitro test according to OECD and GLP guidelines, using the Human Skin Model test.

Two tissues of the human skin model EpiDerm were treated with the test substance for three minutes and one hour, respectively.

50 μL of the liquid test item were applied to each tissue and spread to match the tissue size. Deionised water was used as negative control, 8m KOH was used as positive control.

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 a blue formazan. Formazan production was measured by measuring the optical density (OD) of the resulting solution. After treatment with the negative control, the absorbance values were well above the required acceptability criterion of mean OD > 0.8 for both treatment intervals thus showing the quality of the tissues. The positive control showed clear corrosive effects for both treatment intervals. After three minutes treatment with the test item, the relative absorbance values were reduced to 91.4 %. This value is well above the threshold for corrosion potential (50%). After one hour treatment, relative absorbance values were reduced to 88.7 %. This value, too, is well above the threshold for corrosion potential (15%).

In the guideline, values greater or equal to the threshold are considered as “non-corrosive to skin”. Therefore, the test substance is considered as not corrosive in the Human Skin Model Test.

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

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Recent GLP compliant study, according to international guidelines with detailed test report.

Qualifier:

according to guideline

Guideline:

OECD Guideline 437 (Bovine Corneal Opacity and Permeability Test Method for Identifying Ocular Corrosives and Severe Irritants)

Qualifier:

according to guideline

Guideline:

EU method B.47 (Bovine corneal opacity and permeability test method for identifying ocular corrosives and severe irritants)

GLP compliance:

yes (incl. QA statement)

Species:

other: in vitro

Strain:

other: in vitro

Details on test animals or tissues and environmental conditions:

not applicalbe: in vitro test

Vehicle:

unchanged (no vehicle)

Controls:

yes

Amount / concentration applied:

750 µL of liquid test substance was tested undiluted.

Duration of treatment / exposure:

Exposition time on the corneas was 10 min at 32 ± 1 °C. After thorough rinsing with cMEM with phenol red and final rinsing with cMEM without phenol red, the anterior chamber was filled with cMEM without phenol red, and the corneas were stored for an additional 2 hours at 32 ± 1°C (post-incubation).

Details on study design:

Test vessels:
All vessels used are made of glass or sterilizable plastic. They were sterilised before use by heating to 180 °C (two hours) or autoclavation.
The following vessels were used: Schott-bottles, glass vials, and culture flasks for solutions and media

Negative Control
Sodium chloride solution: 0.9% NaCl (CAS-No. 7647-14-5), dissolved in deionised water.

Positive Control
Dimethyl formamide, DMF, CAS-No. 68-12-2, undiluted

Test System:
Bos primigenius Taurus (Fresh bovine corneas). Fresh bovine eyes were obtained from the slaughterhouse Müller Fleisch GmbH, Enzstr. 2.4, 75217 Birkenfeld, Germany, on the day of the test. The cattle were between 12 and 60 months old. The eyes were transported to the test facility in Hank’s balanced salt solution (supplemented with 0.01% streptomycin and 0.01% penicillin). Then the corneas were dissected and incubated in medium at 32 ± 1 °C in an incubation chamber for 1 hour.

After having carefully cleaned and sterilised the cornea holders, they were kept in the incubation chamber at 32 °C.
On the day of the assay, the MEM without phenol red was supplemented with sodium bicarbonate, L-glutamine and 1% fetal calf serum (= complete MEM) and stored in a water bath at 32 °C ± 1 °C. The same was performed with the MEM with phenol red. After the arrival of the corneas they were examined and only corneas which were free from defects were used. The corneas were excised with a scalpel and cut from the globe with a 2-3 mm ring of sclera around the outside. Each cornea was transferred to a cornea holder in which pre-warmed cMEM without phenol red was filled. The holders were then incubated for one hour in the incubation chamber at 32 °C.

After the initial incubation, the medium was changed and the baseline opacity for each cornea was recorded. None of the corneas showed tissue damage; therefore, all corneas were used. The baseline opacity was measured by placing the holder with the cornea in a spectral photometer and recording the absorption at 570 nm.
For each treatment group (negative control solution, test item and positive control), three replicates were used. After removal of the pre-incubation medium, 750 µl negative control solution resp. test item resp. positive control were applied to each replicate.

Irritation parameter:

in vitro irritation score

Value:

>= 0.623

Vehicle controls validity:

valid

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

no indication of irritation

Irritant / corrosive response data:

Not irritating

Other effects:

The test item FAV ES (FAV BUTILATO) showed no effects on the cornea of the bovine eye.

Table 1: Absorption and Opacity Values Negative Control

Parameter	Negative Control
Absorption before exposition	0.1642	0.1553	0.1469
Absorption after exposition	0.2101	0.1917	0.1901
Opacity before exposition	1.4595	1.4299	1.4025
Opacity after exposition	1.6222	1.5549	1.5492
Opacity Difference	0.1627	0.1250	0.1467

Mean opacity difference of the negative control is 0.1448.

 

Table 2 Absorption and Opacity Values Test Item and Positive Control

Parameter	Test ItemFAV ES (FAV BUTILATO)			Positive Control
Absorption before exposition	0.1455	0.1680	0.1284	0.1276	0.1759	0.1385
Absorption after exposition	0.2263	0.3010	0.3215	1.8965	1.9475	1.7109
Opacity before exposition	1.3980	1.4723	1.3440	1.3415	1.4993	1.3756
Opacity after exposition	1.6838	1.9999	2.0965	78.7952	88.6135	51.3925
Opacity Difference	0.2859	0.5275	0.7525	77.4537	87.1142	50.0169

Table 3 IVIS

Test Group	IVIS	Mean IVIS	Relative Standard Deviation IVIS
Negative Control 0.9% NaCl	0.253	0.187	32.5
	0.133
	0.177
Test Item	0.639	0.623	10.5%
	0.551
	0.678
Positive Control DMF undiluted	92.102	86.251	22.3%
	101.837
	64.815

Interpretation of results:

not irritating

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

The substance resulted not irritating

Executive summary:

The eye damaging potential of the substance was assessed via an in vitro test according to OECD and GLP guidelines, using the BCOP test.

Bovine corneas were used. They were collected from slaughtered cattle which were between 12 and 60 months old.

The test item was brought onto the cornea of a bovine eye which had been incubated with cMEM without phenol red at 32 ± 1 °C for one hour and whose opacity had been measured. The test item was incubated on the cornea for 10 minutes at 32 ± 1 °C. After removal of the test item and two hours post-incubation, opacity and permeability values were measured. Physiological sodium chloride solution was used as negative control. The negative control showed no irritating effect on the cornea. DMF undiluted was used as positive control. The positive control induced a severe irritation on the cornea. The test item showed no effects on the cornea of the bovine eye. The calculated IVIS (in vitro irritancy score) is 0.623.

 

According to OECD Guideline no. 437 (Jul. 2013), a substance with an IVIS ≤ 3 requires no classification for eye irritation or serious eye damage.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Respiratory irritation

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Skin irritation / corrosion

Information on the tendency of the substance to cause skin irritation or corrosion can be obtained from the in vitro skin corrosion test (Andres, 2013), and from the acute dermal toxicity test (Oberto, 2014).

The in vitro skin corrosion test is carried out according to OECD and GLP guidelines, using the Human Skin Model test.

Two tissues of the human skin model EpiDerm were treated with the test substance for three minutes and one hour, respectively.

50 μL of the liquid test item were applied to each tissue and spread to match the tissue size. Deionised water was used as negative control, 8m KOH was used as positive control.

After three minutes treatment with the test item, the relative absorbance values were reduced to 91.4 %. This value is well above the threshold for corrosion potential (50%). After one hour treatment, relative absorbance values were reduced to 88.7 %. This value, too, is well above the threshold for corrosion potential (15%).

These results show that the substance is not corrosive under the conditions of the Human Skin Model Test.

Furthermore, the acute toxicity of the test substance following dermal administration was assessed according to OECD and GLP guidelines. A single dose of 2000 mg/kg was administered to a group of 5 male and 5 female animals for 24 hours. Clinical and necropy examinations did not reveal any abnormalities in the animals nor at the treated site.

Based on this information, it can be concluded that the test substance does not induce effects of skin irritation or corrosion.

Eye irritation / corrosion

Two studies are available in which the tendency of the substance to cause eye irritation or corrosion is assessed. It concerns one in vitro study (Andres, 2013 ) and one in vivo study (Oberto, 2014).

In the in vitro study, the eye dammaging potential of the substance was assessed using the BCOP test according to OECD and GLP guideliens. The test item was brought onto the cornea of a bovine eye which had been incubated with cMEM without phenol red at 32 +/- 1°C for one hour and whose opacity had been measured. The test item was incubated on the cornea for 10 minutes at 32 ± 1 °C. After removal of the test item and two hours post-incubation, opacity and permeability values were measured.Physiological sodium chloride solution was used as negative control. The negative control showed no irritating effect on the cornea.DMF undiluted was used as positive control. The positive control induced a severe irritation on the cornea. The test item showed no effects on the cornea of the bovine eye. The calculated IVIS (in vitro irritancy score) is 0.623.

These results show that the substance does not cause eye irritation or serious eye dammage.

The in vivo eye irritation study is conducted according to OECD and GLP guidelines. A 0.1 mL aliquot of the test item was introduced into the right eye of a total of 3 animals. The resulting reaction to treatment was assessed approximately 1, 24, 48, and 72 hours, 7, 14 and 21 days after dosing. No irritation at either the conjunctivae, iris or cornea was recorded in any treated animal during the whole observation period of the study. There were no signs of pain/distress after dosing. Changes in body weight were not remarkable. There was no indication of a systemic effect related to treatment.

These results indicate that the test item, FAV-ES, has no effect on the eye of the rabbit.

Justification for selection of skin irritation / corrosion endpoint:
This study is a well documented, recent GLP study according to international guidelines.

Justification for selection of eye irritation endpoint:
This study is a well documented, recent GLP study according to international guidelines.

Justification for classification or non-classification

Skin irritation

As no effects of skin irritation or corrosion are observed, the substance is not to be classified according to the criteria described in EU Regulation No. 1272/2008 on the Classification, Labelling and Packaging of Substances and Mixtures (CLP) nor Directive 67/548/EEC (Dangerous Substances Directive).

 

Eye irritation

As no effects of eye irritation or eye dammage are observed, the substance is not to be classified according to the criteria described in EU Regulation No. 1272/2008 on the Classification, Labelling and Packaging of Substances and Mixtures (CLP) norDirective 67/548/EEC (Dangerous Substances Directive).