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Toxicological information

Eye irritation

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Administrative data

eye irritation: in vivo
Type of information:
experimental study
Adequacy of study:
supporting study
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
study well documented, meets generally accepted scientific principles, acceptable for assessment

Data source

Reference Type:
Changes in eye blink frequency as a measure of trigeminal stimulation by exposure to limonene oxidation products, isoprene oxidation products and nitrate radicals
Klenø J and Wolkoff P
Bibliographic source:
Int Arch Occup Environ Health 77: 235–243

Materials and methods

Test guideline
no guideline followed
Principles of method if other than guideline:
One eye of human volunteers was exposed to ozone for 20 minutes. Before, during and after exposure the blink frequency of the exposed eye was calculated.
GLP compliance:

Test material

Constituent 1
Chemical structure
Reference substance name:
EC Number:
EC Name:
Cas Number:
Molecular formula:
Test material form:
Specific details on test material used for the study:
Ozone was generated photochemically by the irradiation of pure oxygen with a thermostat-controlled mercury lamp with a high performance
power supply.

Test animals / tissue source

Details on test animals or tissues and environmental conditions:
Males, non-smoker, absence of any pathological eye history and otherwise healthy. No use of contact lenses for 2 weeks before exposure. The subjects used no systemic medication likely to provoke dry eyes.

Test system

other: air
other: during exposure values were compared with pre- and post-exposure values.
Amount / concentration applied:
71 ppb.
Ozone was monitored continuously in the reaction mixture with a calibrated chemiluminescence monitor (model 265 A, API, San Diego, Calif., USA).
Duration of treatment / exposure:
20 minutes
Observation period (in vivo):
8 minutes before and after exposure
Number of animals or in vitro replicates:
Details on study design:
A specially designed glass eyepiece was utilised to restrict the exposure to the eye. The eyepiece was connected to the reaction flow tube or the stainless-steel chamber via a Teflon tube. A digital video-camera (Sony DCR-PC110E PAL; 25 frames/s) recorded the subject’s blinking; in addition, comments by the subjects about perception were recorded. The temperature was 21°C ± 2°C and the relative humidity was 31% ± 9% in the laboratory. The subjects were exposed single-blind in their non-dominant eye, which sometimes caused minor visual disturbances through the eyepiece. The dominant eye was unaffected. The subjects were neither informed about the nature of stimulus nor could they smell it. An educational video-film was shown during the session to prevent the subjects from cat-napping. The sessions was made up of four successive stages acclimatisation (clean air for 3 min); initial baseline recording (clean air for 8 min) ozone for 20 min and recovery (clean air for 8 min). The shift from pre-exposure to during and during to post-exposure stages involved a manual change of eyepieces and interrupted the stages for ca. 1 min. The subjects were exposed in the morning.

Effects on blinking frequency (BF) were measured. To this aim digital video-camera (Sony DCR-PC110E PAL; 25 frames/s) recorded the subject’s blinking; in addition, comments by the subjects about perception were recorded. All video recordings were critically viewed, and each complete blink was stored. The BF was calculated in sequences of non-overlapping 4 min averages. The data were transformed into a normal distribution by taking the logarithm of the BF. The change of BF caused by exposure to a reaction mixture or single compound was modelled by linear regression analysis. A possible effect of clean air was tested in sham exposed controls.

Results and discussion

In vivo

Remarks on result:
not measured/tested
Other effects:
No change in blinking frequence was noted during exposure when compared with pre- and post-exposure values, while four out of eight subjects reported irriation from this substance.

Applicant's summary and conclusion

Interpretation of results:
study cannot be used for classification
Although part of the subjects reported irritation from ozone at a level of 71 ppb in air this did not result in a measurable change in blinking frequency of the eyes. It is concluded that the ozone concentration was close to a non-irritating level.
Executive summary:

Human eyes were exposed to 71 ppb of ozone for 20 minutes in vivo and blink frequency was measured as a measure of trigeminal stimulation of the eye. Blink frequency during exposure was compared with blink frequency before and after exposure. No effects of the treatment on blink frequency were observed. However, four out of the 8 subjects experienced eye-irritation at this exposure