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Toxicological information

Eye irritation

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Administrative data

Endpoint:
eye irritation: in vivo
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
comparable to guideline study with acceptable restrictions

Data source

Reference
Reference Type:
publication
Title:
Ophthalmic toxicology of dichloromethane
Author:
Ballantyne B, Gazzard MF, Swanson DW
Year:
1976
Bibliographic source:
Toxicol 6, 173-187

Materials and methods

Test guideline
Qualifier:
no guideline followed
Principles of method if other than guideline:
Investigation of DCM effect as a liquid or vapour on rabbit eye, by different methods: instillation of 0.1 ml or 0.01 ml liquid DCM in rabbit eye and 10 min exposure to 1750 or 17500 mg/m3 DCM vapour in 1 m3 exposure chamber.
GLP compliance:
no
Remarks:
did not exist at that time

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
Analar DCM (British Drug Houses Ltd), redistilled before tests, was used.

Test animals / tissue source

Species:
rabbit
Strain:
New Zealand White

Test system

Vehicle:
unchanged (no vehicle)
Controls:
not required
Amount / concentration applied:
- 0.1 ml
- 0.01 ml (simulating splash contamination)
- 1750 mg/m3 (as a vapour)
- 17500 mg/m3 (as a vapour)
Duration of treatment / exposure:
The liquid DCM was instilled once.
The effects of DCM vapour were investigated as 10-min exposures
Observation period (in vivo):
at 10 min, 1, 6, and 24 h, and thereafter daily for 2 weeks
Number of animals or in vitro replicates:
92 animals total
18 for eye irritation studies,
12 for a histopathological evaluation of eye lesions,
24 for in vivo measurement of corneal thickness,
30 for measurement of intraocular tension, and
8 to assess the effect of therapy
Details on study design:
The liquid substance was instilled once into the conjunctival sac of the right eye, after which the lids were held together for a few seconds.
The effects of DCM vapour were investigated as 10-min exposures to 1750 mg/m3 or 17 500 mg/m3. Rabbits were exposed in a 1 m3 chamber using a "Spraying Systems" nozzle to generate the DCM aerosol, and the atmospheric concentration of DCM was continuously monitored by infra-red analysis

30 rabbits had 0.1 ml DCM instilled into the conjunctival sac of the right eye, of which;
6 animals were used for eye irritation studies,
6 for in vivo measurement of corneal thickness,
6 for measurement of intraocular tension, and
12 for a detailed histopathological study.

18 rabbits had 0.01 ml DCM instilled, of which;
6 were used for eye irritation studies,
6 for corneal thickness measurements, and
6 for intraocular tension measurements.

30 rabbits were exposed to 1750 mg/m3 or 17 500 mg/m3 vapour DCM.
From each group of 15 rabbits, 3 were examined for eye irritant effects, 6 used for measurement of corneal thickness, and 6 for measurement of intraocular tension.

Because of technical difficulties encountered in measuring intraocular tension after instilling 0.1 ml DCM, tensions were also measured in a further 6 rabbits receiving 0.02 ml DCM.

In the histopathological study of the effects of 0.1 ml DCM, rabbits were sacrificed by an intravenous overdose of sodium pentobarbitone at 6 h, and at 1, 2, 4, 7 and 14 days. Eyes and eyelids we're removed, fixed in 10% phosphate-buffered neutral formalin, and 6 μm-thick sections of paraffin blocked material stained with haematoxylin and eosin and by the periodic acid-Schiff technique.

8 rabbits were used to assess the effects of treating with a decongestant preparation after contaminating the eye with liquid DCM. The commercially available preparation Vasocon-A was used; this contains an antihistamine, antazoline phosphate (0.5% w/v), and an a-adrenergic sympathomimetic, naphazoline hydrochloride (0.05%). All rabbits had 0.01 ml DCM instilled; of these 4 had no further treatment, but the other 4 had a single drop of Vasocon-A instilled twice daily for 4 days, starting 15 min after contamination with DCM. Eyes were inspected for irritant effects and intraocular tension was measured at 1 and 6 h, and 1, 2, 3, 4 and 7 days.

Subjective assessment of irritant effects was made at 10 min, 1, 6 and 24 h, and thereafter daily for 2 weeks. Particular attention was paid to excess lachrymation, inflammatory changes in the eyelids (blepharitis), injection of conjunctivae, chemosis, sloughing, iritis, keratitis and vascularization of the cornea. Each effect was scored on a system whose guidelines were as follows:
Grade 0 - no effect
Grade 1 - a just detectable effect
Grade 2 - mild effect
Grade 3 - moderate effect
Grade 4 - marked effect
Grade 5 - severe effect with complication
Precise definition of the various grades for the different effects have been described by Ballantyne (1974, 1975). At the ends of the periods of inspection rabbits were sacrificed by an overdose of intravenously injected 6% sodium pentobarbitone, and their eyes removed for histological examination.

Corneal thickness was measured in vivo using an optical depth measuring device attached to a Zeiss slit-lamp microscope. Thickness was measured prior to contamination of the eyes, and then at 30 min, 1 and 6 h, and thereafter daily for up to 11 days. At every period of inspection 5 measurements were taken on each eye and the mean value recorded.

Intraocular tension was measured with a Draeger hand-held applanation tonometer. Control tensions were taken before contamination of the eye and, when chemosis was not so marked as to interfere with applanation measurements, at 10 min, 1, 5 and 24 h, and thereafter daily until tensions returned to normal.


Results and discussion

In vivo

Resultsopen allclose all
Irritation parameter:
other: lachrymation
Basis:
mean
Time point:
other: 10 min, 1h, 6h, and daily up to 14 days
Score:
0 - 2.2
Max. score:
5
Reversibility:
fully reversible within: 8 days
Remarks on result:
other: volume of DCM: 0.1 ml
Irritation parameter:
other: blepharitis
Basis:
mean
Time point:
other: 10 min, 1h, 6h, and daily up to 14 days
Score:
0 - 1.9
Max. score:
5
Remarks on result:
other: volume of DCM: 0.1 ml
Irritation parameter:
other: conjunctival injection
Basis:
mean
Time point:
other: 10 min, 1h, 6h, and daily up to 14 days
Score:
0 - 2.1
Max. score:
5
Remarks on result:
other: volume of DCM: 0.1 ml
Irritation parameter:
chemosis score
Basis:
mean
Time point:
other: 10 min, 1h, 6h, and daily up to 14 days
Score:
0 - 2.8
Max. score:
5
Reversibility:
fully reversible within: 7 days
Remarks on result:
other: volume of DCM: 0.1 ml
Irritation parameter:
other: sloughing
Basis:
mean
Time point:
other: 10 min, 1h, 6h, and daily up to 14 days
Score:
0 - 1.1
Max. score:
5
Reversibility:
fully reversible within: 7 days
Remarks on result:
other: volume of DCM: 0.1 ml
Irritation parameter:
other: iritis
Basis:
mean
Time point:
other: 10 min, 1h, 6h, and daily up to 14 days
Score:
0 - 1.1
Max. score:
5
Reversibility:
fully reversible within: 8 days
Remarks on result:
other: volume of DCM: 0.1 ml
Irritation parameter:
other: keratitis
Basis:
mean
Time point:
other: 10 min, 1h, 6h, and daily up to 14 days
Score:
0 - 1.3
Max. score:
5
Remarks on result:
other: volume of DCM: 0.1 ml
Irritation parameter:
other: corneal vascularization
Basis:
mean
Time point:
other: 10 min, 1h, 6h, and daily up to 14 days
Score:
0 - 0.4
Max. score:
5
Remarks on result:
other: volume of DCM: 0.1 ml
Irritant / corrosive response data:
Flooding the rabbit eye with 0.1 ml liquid DCM caused moderate inflammation of the conjunctiva and eyelids, together with corneal injury. Excess lachrymation and chemosis persisted for about a week, and injection of the conjunctiva and lid margins for nearly 2 weeks. Iritis, occurring in two-thirds of animals, persisted for 1 to 7 days. Keratitis, varying from just detectable to marked, and accompanied by corneal neovascularization also occurred in those with iritis.
With 0.01 ml DCM, more equivalent to splash contamination of the eye, the inflammatory effects on conjunctiva and eyelid were just as marked as those occurring with 0.1 ml, but less persistent.
Other effects:
The potential of DCM to cause corneal injury was confirmed by the in vivo measurements of corneal thickness; a mean peak in-crease of 59% occurred at 6 h with thickness not returning to normal for 8 or 9 days. The histopathological findings of oedema, congestion, neutrophil infiltration, goblet cell proliferation and epithelial detachment of the conjunctiva, with neutrophil infiltration and denudation of the corneal epithelium are typical of chemical injury to the eye. However, both the macroscopic and histological observations demonstrated that healing occurs.
Although macroscopic signs of eye irritation were not apparent after ex¬posure to DCM vapour, small increases in corneal thickness occurred which were related to the degree of exposure.
Contamination of the eye with DCM, both liquid and vapour, caused increases in intraocular tension.

Applicant's summary and conclusion

Interpretation of results:
Category 2 (irritating to eyes) based on GHS criteria