Amines, C12-14-alkyl, C6-10-alkyl phosphates

Eye irritation

Administrative data

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:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2011

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

Deviations:

The following deviations from the study plan were observed:
+ Cuvettes with a pathlength of 0.2 cm were used in the measurement of the Fluorescein-
Na solution in the spectrophotometer and all absorptions were multiplied with
the coefficient 5. The deviation was considered as uncritical since the test results
were not influenced.
+ The positive control benzalkonium chloride solution (5%) was dissolved in demineralised
water instead of 0.9% sodium chloride solution. The deviation can be seen as
uncritical since it was incorrectly stated in the study plan.
The deviations were assessed and signed by the deputy study director on 11. Jan. 2011.

Deviations:

yes

Remarks:

The deviation can be seen as uncritical

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

Amines, C12-14-alkyl, C6-10-alkyl phosphates

EC Number:

271-663-3

EC Name:

Amines, C12-14-alkyl, C6-10-alkyl phosphates

Cas Number:

68603-55-4

Molecular formula:

not available (UVCB)

IUPAC Name:

Amines, C12-14-alkyl, C6-10-alkyl phosphates

Test material form:

liquid: viscous

Test animals / tissue source

Species:

other: bovine eyes

Strain:

not specified

Details on test animals or tissues and environmental conditions:

Freshly 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 with media at 32 ± 1 oc in an incubation chamber for 1 hour.

Test system

Vehicle:

unchanged (no vehicle)

Controls:

yes, concurrent positive control

yes, concurrent negative control

Amount / concentration applied:

in average, 214.3 mg of the test item were tested neat and applied directly on the cornea using a weight board. The test item was given on the epithelium in such a manner that as much as possible of the corneawas covered with test item. The test item is a surface active liquid. Surfactants (solids or liquids) are usually tested at a concentration of 10% in a 0.9% sodium chloride solution, deionised water or other solvents which have no adverse effects in the test system. The test item Deophos 228 was not soluble in one of the solvents mentioned above in the demanded concentration (10%). Since no solutionwas feasible, the test item was applied directly on the cornea without dilution or preparation of a solution.

Duration of treatment / exposure:

10 minutes

Duration of post- treatment incubation (in vitro):

90 ± 5 min

Number of animals or in vitro replicates:

3

Results and discussion

In vitro

Results

Irritation parameter:

in vitro irritation score

Run / experiment:

mean

Value:

ca. 0.785

Vehicle controls validity:

not specified

Negative controls validity:

valid

Positive controls validity:

valid

Any other information on results incl. tables

IVIS was calculated using the values in tables 9.1-a and 9. I-b and the equation stated in chapter 8.3. Example: IVIS (Deophos 228, Repl. 1) = (0.0893 - 0.2416) + [15 * (5 * 0.0142 - 0.0200)] = 0.6127

The calculated IVIS for each replicate and the corresponding means are presented in the following table:

Table 9.2.a IVIS

			Relative Standard Deviation IVIS
Negative Control 0,9% NaCl	0.6757	0.5420	21.4%
	0.4836
	0.4671
Deophos 228	0.6127	0.7850
	1 .5786
	O. 1644
Positive Control 5% Benzalkonium chloride solution	88.5716	108.6360	17.0%
	112.4971
	124.8378

Note: the high relative standard deviation of the IVIS of test item is due to mathematical reasons, as the respective mean is very small.

Parameter                                               Criterion

IVIS of ne ative control 0.9% NaCl	0 - 3	Found 0.5420	Assessment ok
IVIS of positive control 5% Benzalkonium chloride solution	55.4 —	108.6360	Too high

Values for negative control were within the range of historical data of the test facility (see 14, page 20). Therefore, the test system was acceptable. The values for positive control were not within the range of historical data of the test facility. This can be seen as uncritical because the range of the historical data was quite narrow since it referred only to six individual replicates. Besides that, the high value of the positive control can be seen as uncritical because of the fact that it showed clearly very severe eye irritating effects which confirms that the test system is acceptable.

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

This in vitro study was performed to assess the corneal irritation and darnage potential of Deophos 228 by quantitative measurements of changes in opacity and permeability in a bovine cornea. The test item Deophos 228 was brought onto the cornea of a bovine eye which previously had been incubated with cMEM without Phenol red at 32 ± 1°C for one hour and whose opacity had been determined. The lest item was incubated on the cornea for 10 minutes at 32 ± 1°C. After removal of the lest item and two hours post-incubation, opacity and permeability values were measured. Physiological sodium chloride solution was used as negative control; benzalkonium chloride (5% solution in demineralised water) was used as positive control. The positive control induced a very severe irritation on the cornea, mean IVIS was 108.6360. The negative control showed no irritation, mean IVIS was 0.5420. The lest item was tested pure. A mean IVIS of 0.7850 was calculated, corresponding to a classification as not eye irritant. No observations were made which might cause doubts concerning the validity of the study outcome. The lest is considered valid.

Executive summary:

One valid experiment was performed. 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. 5% benzalkonium chloride solution was used as positive control. The positive control induced a very 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. 7850. ln conclusion, it can be stated that in this study and under the experimental conditions reported, the test item possesses no eye irritation potential.