Alcohols, C16-18 and ethoxylated C16-18 , phosphates (20 moles ethoxylation)

Eye irritation

Administrative data

Endpoint:

eye irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

July 04, 2017

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2017

Report date:

2017

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 437 (Bovine Corneal Opacity and Permeability Test Method for Identifying i) Chemicals Inducing Serious Eye Damage and ii) Chemicals Not Requiring Classification for Eye Irritation or Serious Eye Damage)

Deviations:

yes

Remarks:

Deviation was considered to have not affected the integrity or validity of the study.

Test guideline 2

Qualifier:

according to guideline

Guideline:

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

Deviations:

yes

Remarks:

Deviation was considered to have not affected the integrity or validity of the study.

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

Alcohols, C16-18 and ethoxylated C16-18 , phosphates (20 moles ethoxylation)

Molecular formula:

C18H39O4P1 (monoester representative, i.e., mono- C18 PSE) C56H115O24P1 (ethoxylated monoester representative, i.e., mono- C16 AE20 PSE) C32H67O4P1 (diester representative, i.e., di- C18 PSE) C112H227O44P1 (ethoxylated diester representative, i.e., di- C16 AE20 PSE)

IUPAC Name:

Alcohols, C16-18 and ethoxylated C16-18 , phosphates (20 moles ethoxylation)

Test material form:

solid

Test animals / tissue source

Species:

cattle

Strain:

not specified

Details on test animals or tissues and environmental conditions:

Source of Bovine Eyes
Eyes from adult cattle (typically 12 to 60 months old) were obtained from a local abattoir as a by-product from freshly slaughtered animals. The eyes were excised by an abattoir employee after slaughter, and were placed in Hanks’ Balanced Salt Solution (HBSS) supplemented with antibiotics (penicillin at 100 IU/mL and streptomycin at 100 µg/mL). They were transported to the test facility over ice packs on the same day of slaughter. The corneas were prepared immediately on arrival.

Test system

Vehicle:

physiological saline

Controls:

yes, concurrent vehicle

yes, concurrent positive control

yes, concurrent negative control

Amount / concentration applied:

0.75 mL of the 20% w/v test substance solution in sodium chloride 0.9% w/v or control substances (sodium chloride 0.9% w/v as negative control, 20% w/v imidazole solution in sodium chloride 0.9% w/v as positive control)

Duration of treatment / exposure:

240 minutes

Duration of post- treatment incubation (in vitro):

32 ± 1ºC for 90 minutes

Number of animals or in vitro replicates:

Three replicates per substance

Results and discussion

In vitro

Results

Irritation parameter:

in vitro irritation score

Run / experiment:

Test substance

Value:

ca. 89.1

Vehicle controls validity:

valid

Negative controls validity:

valid

Positive controls validity:

not valid

Remarks:

The deviation was considered to have not affected the integrity or validity of the study

Remarks on result:

positive indication of irritation

Remarks:

Category 1 (irreversible effects on the eye) based on EU CLP criteria

Other effects / acceptance of results:

The positive control group had an overall IVIS of 127.0, which was marginally higher than the criteria range set for an acceptable test. However, as the score was only marginally exceeded, it was decided that this result was acceptable as the positive control group was still providing its intended function which is to show the sensitivity of the test system to a known ocular irritant. This deviation was considered to have not affected the integrity or validity of the study. The negative control gave opacity of ≤2.4 and permeability ≤0.072. The negative control acceptance criteria were therefore satisfied.

Any other information on results incl. tables

Results

Corneal Opacity and Permeability Measurement

Individual and mean corneal opacity measurements and individual and mean corneal permeability measurements are given in below table:

Table1: Individual and Mean Corneal Opacity and Permeability Measurements

Treatment	Cornea Number	Opacity				Permeability Optical Density (OD)		In Vitro Irritancy Score
		Pre-Treatment	Post-Treatment	Post-Treatment-Pre‑Treatment	Corrected Value		Corrected Value
Negative Control #	1	3	4	1		0.012
	2	3	4	1		0.000
	3	2	4	2		0.000
	Mean			1.3		0.004		1.4
Positive Control #	4	2	105	103	101.7	3.965	3.961
	5	2	88	86	84.7	1.895	1.891
	6	2	83	81	79.7	1.820	1.816
	Mean				88.7		2.556	127.0
Test Substance	7	3	94	91	89.7	0.504	0.500
	8	2	85	83	81.7	0.402	0.398
	9	2	79	77	75.7	0.468	0.464
	Mean				82.3		0.454	89.1

#= Control data shared with Envigo - Shardlow study number LM55TK and XL29CC

 

Corneal Epithelium Condition

The condition of each cornea is given in below table:

Table 2: Corneal Epithelium Condition Post Treatment

Treatment	Cornea Number	Observation Post Treatment
Negative Control #	1	Clear
	2	Clear
	3	Clear
Positive Control #	4	Cloudy
	5	Cloudy
	6	Cloudy
Test Substance	7	Cloudy
	8	Cloudy
	9	Cloudy

#= Control data shared with Envigo - Shardlow study number LM55TK and XL29CC

 The corneas treated with the test substance were cloudy post treatment. The corneas treated with the negative control substance were clear post treatment. The corneas treated with the positive control substance were cloudy post treatment.

 

In Vitro Irritancy Score

The In Vitro irritancy scores are summarized as follows:

Treatment	In Vitro Irritancy Score
Test Substance	89.1
Negative Control	1.4
Positive Control	127.0

 

Criteria for an Acceptable Test

The positive control In Vitro Irritancy Score was above the range of 65.1 to 123.3. The positive control acceptance criterion was therefore not satisfied. This is reported as a deviation. The negative control gave opacity of ≤2.4 and permeability ≤0.072. The negative control acceptance criteria were therefore satisfied.

 

Conclusion

Based on the study results, the test substance was classified as Category 1 (irreversible effects on the eye) based on GHS criteria

Applicant's summary and conclusion

Interpretation of results:

other: Category 1 (irreversible effects on the eye) based on EU CLP criteria

Conclusions:

Under the study conditions, the test substance was determined as inducing serious eye damage and classified as Category 1 (irreversible effects on the eye) based on EU CLP criteria.

Executive summary:

An in vitro study was conducted to determine the eye irritation potential of the the test substance, ‘mono- and di- C16-18 PSE and C16-18 AE20 PSE’, using the Bovine Corneal Opacity and Permeability (BCOP) method, according to the OECD Guideline 437 and EU Method B.47, in compliance with GLP. The test substance was applied to test system at a concentration of 20% w/v in 0.9% w/v sodium chloride for 240 minutes followed by post exposure period at 32 ± 1ºC for 90 minutes. Negative and positive control substances were tested concurrently. The two endpoints, decreased light transmission through the cornea (opacity) and increased passage of sodium fluorescein dye through the cornea (permeability) were combined in an empirically derived formula to generate an In Vitro Irritancy Score (IVIS). The test substance IVIS was determined to be 89.1, which is well above the corrosive limit of 55. Therefore, the test substance was classified as Category 1 (irreversible effects on the eye) based on GHS/EU CLP criteria. The positive control IVIS was 127, which was outside the range of 65.1 to 123.3, however, as the score was only marginally exceeded, study author decided that this result was acceptable as the positive control group was still providing its intended function which is to show the sensitivity of the test system to a known ocular irritant. Therefore, this deviation was considered to have not affected the integrity or validity of the study. The negative control gave opacity of ≤1.3 and permeability ≤0.004, therefore the negative control acceptance criteria were satisfied. The test was considered to pass all the validity criterias. Under study conditions, the test substance was considered to be corrosive and classified as Eye Damage 1 (causes serious eye damage) based on EU CLP criteria (Envigo, 2017).