BMS 233110-01

Endpoint summary

Description of key information

Skin irritation potential of the substance BMS 233110-01 was tested according to EU Test Method B4, Directive 92/69/EEC. 500 mg of the test substance were diluted in distilled water and applied semi-occlusively on 3 New Zealand White rabbits. The test material produced a primary irritation index of 0.0 (according to the Draize classification scheme) and no corrosive effects were noted. Therefore, it does not meet the criteria for classification as irritant or corrosive according to the CLP Regulation.

 

Potential of the substance BMS 233110-01 for causing eye damage or irritation was tested according to EU Test Method B5, Directive 92/69/EEC. The test substance was applied to 3 New Zealand White rabbits. Moderate to high conjunctivae redness was observed in all 3 rabbits (1.66 average score). Mild Iridial inflammation in one treated eye was observed over the 72 observation period, but it was resolved by the 7 days. Moderate chemosis of the conjunctivae were observed in all 3 animals. Corneal alterations were observed were observed in 1 animal but was fully reversible in 7 days. As a result, the substance is classified as Eye Irritation Cat 2.

Key value for chemical safety assessment

Skin irritation / corrosion

Link to relevant study records

Reference

Endpoint:

skin irritation / corrosion, other

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2nd Dec - 5th Dec 1998

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

test procedure in accordance with national standard methods

Qualifier:

according to guideline

Guideline:

EU Method B.4 (Acute Toxicity: Dermal Irritation / Corrosion)

GLP compliance:

yes

Specific details on test material used for the study:

Identity of test material same as for substance defined in section 1

Species:

rabbit

Strain:

New Zealand White

Type of coverage:

semiocclusive

Vehicle:

other: The substance was moistened with water and applied to the animal.

Amount / concentration applied:

500 mg

Duration of treatment / exposure:

4 h

Number of animals:

3

Irritation parameter:

erythema score

Basis:

animal #1

Time point:

24/48/72 h

Score:

0

Max. score:

0

Irritation parameter:

erythema score

Basis:

animal #2

Time point:

24/48/72 h

Score:

0

Max. score:

0

Irritation parameter:

erythema score

Basis:

animal #3

Time point:

24/48/72 h

Score:

0

Max. score:

0

Irritation parameter:

edema score

Basis:

animal #1

Time point:

24/48/72 h

Score:

0

Max. score:

0

Irritation parameter:

edema score

Basis:

animal #2

Time point:

24/48/72 h

Score:

0

Max. score:

0

Irritation parameter:

edema score

Basis:

animal #3

Time point:

24/48/72 h

Score:

0

Max. score:

0

Other effects:

There was no sign of dermal effects.

Interpretation of results:

other:

Conclusions:

The test material produced a primary irritiation index of 0.0 to rabbit skin according to the Draize classification scheme. No corrosive effects were noted.

Executive summary:

.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Eye irritation

Link to relevant study records

Reference

Reference 1

Endpoint:

eye irritation, other

Remarks:

in-vivo study

Type of information:

experimental study

Adequacy of study:

key study

Study period:

10th Dec - 22nd Dec 1998

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

test procedure in accordance with national standard methods

Justification for type of information:

An in-vivo eye irriitation study was conducted prior to the ammendments and requirements for Annex VII and the acceptance of the in-vitro BCOP study as part of the testing requirements

Qualifier:

according to guideline

Guideline:

EU Method B.5 (Acute Toxicity: Eye Irritation / Corrosion)

GLP compliance:

yes

Specific details on test material used for the study:

Identity of test material same as for substance defined in section 1

Species:

rabbit

Strain:

New Zealand White

Amount / concentration applied:

ca 50mg

Number of animals or in vitro replicates:

3

Irritation parameter:

conjunctivae score

Remarks:

redness

Basis:

animal #1

Time point:

24/48/72 h

Score:

1.66

Max. score:

2

Reversibility:

fully reversible within: 7 days

Irritation parameter:

conjunctivae score

Remarks:

redness

Basis:

animal #2

Time point:

24/48/72 h

Score:

1.66

Max. score:

2

Reversibility:

fully reversible within: 7 days

Irritation parameter:

conjunctivae score

Remarks:

redness

Basis:

animal #3

Time point:

24/48/72 h

Score:

1.66

Max. score:

2

Reversibility:

fully reversible within: 7 days

Irritation parameter:

chemosis score

Basis:

animal #1

Time point:

24/48/72 h

Score:

1

Max. score:

2

Reversibility:

fully reversible within: 72h

Irritation parameter:

chemosis score

Basis:

animal #2

Time point:

24/48/72 h

Score:

0.66

Max. score:

1

Reversibility:

fully reversible within: 72hrs

Irritation parameter:

chemosis score

Basis:

animal #3

Time point:

24/48/72 h

Score:

0.66

Max. score:

1

Reversibility:

fully reversible within: 72 hrs

Irritation parameter:

cornea opacity score

Basis:

animal #1

Time point:

24/48/72 h

Score:

0.66

Max. score:

1

Reversibility:

fully reversible within: 72h

Irritation parameter:

cornea opacity score

Basis:

animal #2

Time point:

24/48/72 h

Score:

0

Max. score:

0

Reversibility:

fully reversible

Irritation parameter:

cornea opacity score

Basis:

animal #3

Time point:

24/48/72 h

Score:

0

Max. score:

0

Reversibility:

fully reversible

Irritation parameter:

iris score

Basis:

animal #1

Time point:

24/48/72 h

Score:

0.33

Max. score:

1

Reversibility:

fully reversible within: 48h

Irritation parameter:

iris score

Basis:

animal #2

Time point:

24/48/72 h

Score:

0

Max. score:

0

Reversibility:

fully reversible

Irritation parameter:

iris score

Basis:

animal #3

Time point:

24/48/72 h

Score:

0

Max. score:

0

Reversibility:

fully reversible

Irritant / corrosive response data:

Reversibility of any observed effect: Changes fully reversible within 7 days

Other effects:

Discharge and reddening was observed within the first hour after dosing. The redness was still present in all treated eyes at 72 hours but the eyes had recovered by the 7 day observation point. Residual test material was noted around two treated eyes one hour after treatment.

Interpretation of results:

GHS criteria not met

Conclusions:

The test material did not meet the criteria for irritant. However as a precautionary measure the substance is being classified as Irritating to the Eye Cat 2.

Reference 2

Endpoint:

eye irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

17 June to 24 June 1998

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

comparable to guideline study

Justification for type of information:

This study was conducted prior to adoption of the BCOP OECD study and implementation of the Annex VII requirement. The in vitro study was conducted as supporting weight of evidence

Qualifier:

equivalent or similar to guideline

Guideline:

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

Version / remarks:

Study was conducted prior to the adoption and introduction of the BCOP 437 study.

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

White Powder
Stored at room temperature and protected form exposure to light

Species:

cattle

Strain:

not specified

Details on test animals or tissues and environmental conditions:

Bovine eyes were obtained from a local abattoir as a by-product from freshly slaughtered animalsand corneas were used within 24 hours. The eyes were transported in Hanks' Balanced Salt Solution (HBSS), supplemented with Penicillin/Streptomycin, and transported to the lab on ice packs.

Vehicle:

physiological saline

Controls:

yes, concurrent positive control

yes, concurrent negative control

Amount / concentration applied:

750 ul per cornea of the test article suspension or positive control was used

Duration of treatment / exposure:

4 hours

Duration of post- treatment incubation (in vitro):

An opacity measurement was performed immediately (no further incubation) using an opacitometer to determine the change in opacity for each cornea. Permeability of the corneas was determined after a 90 minute incubation period in the presence of sodium fluorescien solution.

Number of animals or in vitro replicates:

Total of 20 corneas but only 9 were suitable for test. 4 corneas treated with test material, 3 treated with negative control, and 2 treated with positive control.

Details on study design:

Preparation of Corneas:
The eyes were grossly examined for damage and those exhibiting defects were discarded. The tissue surrounding the eyeball was carefully pulled away and the cornea was excised such that a 2 to 3 mm rim of sclera was present around the cornea. The isolated corneas were then stored in a petri dish containing HBSS until they were mounted in a corneal holder. The corneas were mounted in the holders with the endothelial side against the O-ring of the posterior half of the holder. The anterior half of the holder was then positioned on top of the cornea and the screws were tightened. Starting with the posterior compartment, the two compartments of the corneal holder were then filled with Minimum Essential Medium Eagle (MEM) without phenol red, with 1% fetal bovine serum (complete MEM). The corneal holders were incubated at 32 +/- 1 degrees C for a minimum of one hour.

Test Article Preparation and pH Determination:
The solid test article, BMS 233110-01, was diluted in USP 0.9% saline (Baxter). An aliquot of the test article was weighed in a glass scintillation vial. Saline was added to achieve a 20% mixture and the vial was vortexed for approximately one minute. The test article was diluted on a weight: volume basis (i.e. mg/ml = 20%).

The pH of the 20% dilution of the test article in saline was determined using pH paper. The prepared dilution was added to pH paper with a 4-7 pH range (EM Science) with 0.2 - 0.5 pH unit increments.

Bovine Corneal Opacity and Permeability Assay:
After a minimum of one hour of incubation, the corneal holders with the corneas in place were removed from the incubator and the medium was removed from both compartments and replaced with fresh medium. The opacity was determined for each cornea using a Spectro Designs OP-KIT opacitometer. Three corneas whose opacity readings were close to the median opacity for all the corneas were selected as the negative control corneas. The medium was then removed from the anterior part of the holder and replaced with either the test compound or positive control. For the negative control corneas, the medium was replaced with fresh complete MEM.

Method for Testing Solid Materials:
The solid test article, BMS 233110-01, was tested as a 20% suspension in saline. An aliquot of seven hundred and fifty ul of the test article suspension or positive control was introduced into the anterior compartment of the holder while slightly rotating the holder (with the corneas maintained in a horizonal position) to ensure uniform distribution of the test article or positive control over the cornea. The corneas were incubated in the presence of either the test article, positive control or negative control at 32 +/- 1 degrees C for approximately 4 hours. After the four hour incubation, the test article or positive control treatments were removed and the epithelial side of the corneas were washed at least 3 times with complete MEM (with phenol red) to ensure complete removal of the test article or positive control. Once the test article and the positive control were removed, the corneas were given a final rinse with complete MEM (without phenol red). The anterior and the posterior compartments were then refilled with fresh complete MEM. The negative control corneas were refed with fresh complete MEM after the four hour incubation. An opacity measurement was performed immediately (no further incubation) using an opacitometer to determine the change in opacity for each cornea.

After the final opacity measurement was performed, the medium was removed from both chambers of the holder. The posterior compartment was refilled with fresh complete MEM. One ml of a 5 mg/ml fluorescein solution was added to the anterior compartment. The corneas were then incubated in a horizontal position, anterior side up for approximately 90 minutes at 32 +/- 1 degrees C. An aliquot of medium was removed from the posterior chamber and placed into tubes numbered corresponding to chamber number. Three hundred and sixty microliters were placed into designated wells of a 96-well plate. The optical density at 490 nm (OD490) was determined using a Molecular Devices Vmax kinetic microplate reader. If the OD490 of a test or control sample was greater than 1.500, a 1:5 dilution of the sample in MEM was made. Samples of the 1:5 dilution were transferred to specified wells on the 96-well plate. The plate was reread and the final reading was saved to a designated print file.

Irritation parameter:

in vitro irritation score

Value:

-1.6

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

no indication of irritation

Irritation parameter:

cornea opacity score

Value:

-1.6

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

no indication of irritation

Irritation parameter:

other: permeability measurement

Value:

-0.001

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

no indication of irritation

Other effects / acceptance of results:

Positive control results: opacity measurement 68.7, permeability measurement 2.120, and in vitro score 100.5

Opacity Measurement = The change in opacity for each cornea was calculated by subtracting the pre-treatment opacity readings from the final opacity readings. The corrected opacity value of each cornea was calculated by subtracting the average change in opacity of the negative control corneas from that of each treated cornea. The mean opacity value of each treatment group was calculated by averaging the mean corrected opacity values of the treated corneas for each treament condition.

Permeability Measurement = The corrected OD490 was calculated by subtracting the mean OD490 of the negative control corneas from the OD490 value of each treated cornea. The mean OD490 value of each treatment group was calculated by averaging the corrected OD490 values of the treated corneas for that treatment condition.

In Vitro Score = Mean Opacity Value + (15 x Mean OD490 Value)

In vitro score:
from 0 to 25 = mild irritant
from 25.1 to 55 = moderate irritant
from 55.1 and above = severe irritant

Criteria for Determination of a Valid Test:
The BCOP assay was accepted when the positive control, imidazole, caused an in vitro score that fell within two standard deviations of the historical mean.

Interpretation of results:

GHS criteria not met

Conclusions:

Based on the opacity measurement of -1.6, permeability measurement of -0.001, and in vitro score of -1.6, this study validly concluded that the test material did not result in eye irritation in cattle. Since the results of the positive control fell within two standard deviations of the historical mean (within a range of 79.8 to 145.0), the assay was considered valid.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed (irritating)

Respiratory irritation

Endpoint conclusion

Endpoint conclusion:

no study available