Singlet oxygen (Δg)-mediated oxidation of cellular and subcellular components: ESR and AFM assays

We report a comprehensive in vitro study of the photo-oxidative stress on different biomolecular and cellular targets generated in the presence of fullerol C60(OH)n , a novel, fullerene-based and water-soluble sensitizer of singlet oxygen (1 g). The photodynamic efficiency of fullerol C60(OH)n was checked by using a singlet oxygen scavenger, TMP-OH, and the electron spin resonance (ESR) technique, which was capable of detecting the resulting paramagnetic product, TEMPOL. ESR was also used to monitor the conformation changes occurring in the spin-labelled protein, T4L lysozyme, which was exposed to the photo-oxidative stress in solutions containing fullerol C60(OH)n. Finally, atomic force microscopy (AFM) experiments were performed to monitor changes in the local elastic properties of living and glutaraldehyde-fixed cells (neurons) exposed to the toxic action of 1 g generated in the presence of fullerol C60(OH)n . Remarkably, the Young’s modulus values measured for both living and fixed neurons revealed a pronounced drop as a function of exposure to the toxic action of 1 g. Thus, our ESR and AFM results bring evidence that the multi-hydroxylated fullerene is an efficient 1 g-generator in aqueous media and might be implemented as a photosensitizer for performing oxidations in biological systems. 6 Author to whom any correspondence should be addressed. Fax: +41(0)21 693 4470, tel: +41(0)21 693 4304. 0953-8984/05/181471+12$30.00 © 2005 IOP Publishing Ltd Printed in the UK S1471 S1472 B Vileno et al