Reaction products of tetraethylenepentamine and maleic anhydride

Eye irritation

Administrative data

Endpoint:

eye irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2015-01-13 to 2015-01-15

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Well documented study performed according to OECD 437 guideline, in compliance with GLP. No deviations were recorded.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2015

Report date:

2015

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)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Test material form:

liquid: viscous

Details on test material:

- Name of test material (as cited in study report): XTA 823- Substance type: yellow slightly viscous liquid- Physical state: liquid- Purity: UVCB (reaction product of tetraethylene pentamine and maleic anhydride 30-60%)- Purity test date: no data- Lot/batch No.: BLW0010057- Expiration date of the lot/batch: 2015-12-01- Stability under test conditions: no data- Storage condition of test material: at approximately 4°C in the dark

Test animals / tissue source

Species:

cattle

Strain:

not specified

Details on test animals or tissues and environmental conditions:

SOURCE OF COLLECTED EYES- Source:bovine eyes from adult cattle were obtained from a local abattoir as a by-product from freshly slaughtered animals- Number of animals: no data- Characteristics of donor animals (e.g. age, sex, weight): adult cattle typically 12 to 60 months old- Storage, temperature and transport conditions of ocular tissue (e.g. transport time, transport media and temperature, and other conditions):The eyes were excised by an abattoir employee after slaughter and 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.- Time interval prior to initiating testing:- indication of any existing defects or lesions in ocular tissue samples: no- Indication of any antibiotics used: noPREPARATION OF CORNEAS- All eyes were macroscopically examined before and after dissection. Only corneas free of damage were used. - The cornea from each of selected eye was removed leaving a 2 to 3 mm rim of sclera to facilitate handling. The iris and lens were peeled away from the cornea. The isolated corneas were immersed (epithelial side uppermost) in a dish containing HBSS until the were mounted in Bovine Corneal Opacity and Permeability (BCOP) holders.- The anterior and posterior chambers of each BCOP holder were filled with complete Eagle's minimum essential medium (MEM) and plugged. The holders were incubated at 32 +/- 1°C for 60 minutes. At the end of the incubation period each cornea was examined for defects. Only corneas free of damage were used.PRE-TREATMENT OPACITY READING- A pre-treatment opacity reading was taken for each cornea using a calibrated opacitometer. The average opacity for all corneas was calculated.

Test system

Vehicle:

unchanged (no vehicle)

Controls:

yes

Amount / concentration applied:

TEST MATERIAL- Amount(s) applied (volume or weight with unit): 0.75 ml, used as suppliedNEGATIVE CONTROL: 0.9% w/v sodium chloride solution- Amount(s) applied (volume or weight with unit): 0.75 ml- Lot/batch no. (if required): 301038501- Purity: 0.9%- Expiry date: 2015-09-01- Storage conditions: at room temperaturePOSITIVE CONTROL: ethanol- Amount(s) applied (volume or weight with unit): 0.75 ml- Lot/batch no. (if required): STBD7546V- Purity: >99.8%- Expiry date: 2015-08-11- Storage conditions: at room temperature

Duration of treatment / exposure:

10 minutes

Duration of post- treatment incubation (in vitro):

220 minutes

Number of animals or in vitro replicates:

3 eyes/group; 3 groups

Details on study design:

REMOVAL OF TEST SUBSTANCE- Washing (if done): at the end of the exposure period the test item and control items were removed from the anterior chamber and the cornea was rinsed 3 times with fresh complete minimum essential medium (MEM) containing phenol red before a final rinse with complete MEM. The anterior chamber was refilled with fresh complete MEM.- Time after start of exposure: 10 minutes- After a post-treatment opacity reading and visual observation of each cornea the holders were incubated at 32 +/- 1°C for 120 minutes. After incubation the holders were removed from the incubator and a final opacity reading was taken. Each cornea was visually observed.APPLICATION OF SODIUM FLUORESCEIN- Following the opacity measurement the permeability of the corneas to sodium fluorescein was evaluated. The medium from the anterior chamber was removed and replaced with 1 ml of sodium fluorescein solution (4 mg/ml). The dosing holes were plugged and the holders incubated, anterior chamber uppermost, at 32 +/- 1°C for 90 minutes.PERMEABILITY DETERMINATIONS- After incubation the medium in the posterior chamber of each holder was decanted and retained.- 360 µl of medium representing each cornea was applied to a designated well on a 96-well plate and the optical density at 492 mm (OD492) was measured using the Anthos 2001 microplate reader.SCORING SYSTEM:- The In Vitro Irritancy Score was calculated using the following formula:In Vitro Irritancy Score = mean opacity value + (15 x mean OD492 value)- Additionally, the opacity and permeability values were evaluated independently to determine wether the test item induced a response through only one of the two endpoints. - The test item was classified according to the prediction model below:IVIS Classification<=3 No category. Not requiring classification to UN GHS or EU CLP>3; <=55 No prediction of eye irritation can be made>55 Category 1. UN GHS or EU CLP causes serious eye damage

Results and discussion

In vitro

Results

Results 1

Irritation parameter:

in vitro irritation score

Run / experiment:

test item

Value:

0.9

Vehicle controls validity:

not examined

Negative controls validity:

valid

Positive controls validity:

valid

Results 2

Irritation parameter:

in vitro irritation score

Run / experiment:

negative control

Value:

3.4

Results 3

Irritation parameter:

in vitro irritation score

Run / experiment:

positive control

Value:

40.9

Other effects / acceptance of results:

OTHER EFFECTS:- Visible damage on test system: Corneal epithelium condition: the corneas treated with the test item were clear post treatment and post incubation. The corneas treated with te negative control item were clear post treatment and post incubation. The corneas treated with the positive control item were cloudy post treatment and post incubation.DEMONSTRATION OF TECHNICAL PROFICIENCY: no dataACCEPTANCE OF RESULTS: - Acceptance criteria met for negative control: negative control gave opacity of <=4.7 and permeability <=0.080. The negative control acceptance criteria were therefore satisfied- Acceptance criteria met for positive control: positive control IVIS was within the range of 27.8-51.0. The positive control acceptance criterion was therefore satisfied- Range of historical values if different from the ones specified in the test guideline: no data

Applicant's summary and conclusion

Interpretation of results:

not classified

Remarks:

Criteria used for interpretation of results: EU

Conclusions:

Based on the results (IVIS score < 3), the test substance is considered not irritant to eyes and should not be classified for eye irritation.