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ToxSci Advance Access published online on February 18, 2003
Toxicological Sciences, doi:10.1093/toxsci/kfg014
Toxicological Sciences © Society of Toxicology 2003; all rights reserved
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Received August 20, 2002; accepted December 10, 2002
© 2003 Society of Toxicology

In Vitro Toxicology and Alternative Testing

Resazurin Metabolism Assay Is a New Sensitive Alternative Test in Isolated Pig Cornea

Sébastien Perrot 1*, Hélène Dutertre-Catella 2, Chantal Martin 3, Patrice Rat 1, Jean-Michel Warnet 1

1 Laboratoire de Toxicologie, Faculté des Sciences Pharmaceutiques et Biologiques, Université Paris V, 75270 Paris cedex 06, France; Laboratoire de Toxicologie, Faculté des Sciences Pharmaceutiques, Université de Tours, 37200 Tours, France; Unité de Pharmaco-Toxicologie Cellulaire, Centre Hospitalier National d'Ophtalmologie des XV-XX, 75571 Paris cedex 12, France
2 Laboratoire de Toxicologie, Faculté des Sciences Pharmaceutiques et Biologiques, Université Paris V, 75270 Paris cedex 06, France; Laboratoire de Toxicologie, Faculté des Sciences Pharmaceutiques, Université de Tours, 37200 Tours, France
3 Laboratoire de Toxicologie, Faculté des Sciences Pharmaceutiques et Biologiques, Université Paris V, 75270 Paris cedex 06, France

* To whom correspondence should be addressed. E-mail: sebastien.perrot{at}libertysurf.fr.


  Abstract

The main object of our study was to investigate whether the resazurin metabolism assay is a sensitive surfactant- and alcohol-toxicity test in isolated pig cornea and to compare this recently developed fluorimetric assay with the data collected in the eye irritation reference chemical data bank. Resazurin is a substrate which changes colour in response to metabolic activity. Isolated pig corneas were immersed for 10 minutes in surfactants and alcohols irritant solutions. After incubation, resorufin fluorescence was read and corneal viability was assessed. This corneal viability was compared with the maximal modified average score published in the report of ECETOC. This assay highlighted different concentration- dependent irritation potential of the three surfactants tested and same results were obtained with corneas treated with the alcohols. We observed that the degree of surfactant- and alcohol-induced decrease in corneal viability using the resazurin reduction test was correlated with the in vivo irritancy measurements as determined by the Draize test and scored with the MMAS. This assay allowed to classify the ocular irritancy of the tested surfactants and alcohols in the same ranking order as the Draize classification. Corneal viability measurement can be used as a potential alternative for the toxicological assessment of surfactants and alcohols. The non-toxic, non-radioactive resazurin metabolism assay allows rapid assessment of many samples, with simple equipment and at reduced cost for continuous monitoring of corneal viability. This assay seems to be suitable as a toxicologic screening test for eye irritation determination.

isolated corneal model, toxicological assessment, resazurin, surfactants, alcohols .


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