Singlet oxygen (1Δg)-mediated oxidation of cellular and subcellular components: ESR and AFM assays

Abstract

We report a comprehensive in vitro study of the photo-oxidative stress ondifferent biomolecular and cellular targets generated in the presence of fullerolC60(OH)n, a novel, fullerene-based and water-soluble sensitizer of singlet oxygen(1Δg). The photodynamicefficiency 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. ESRwas also used to monitor the conformation changes occurring in the spin-labelled protein, T4Llysozyme, which was exposed to the photo-oxidative stress in solutions containing fullerolC60(OH)n.Finally, atomic force microscopy (AFM) experiments were performed to monitor changes in the localelastic properties of living and glutaraldehyde-fixed cells (neurons) exposed to the toxic action of1Δg generated in thepresence of fullerol C60(OH)n. Remarkably, the Young’s modulus values measured for both living and fixedneurons revealed a pronounced drop as a function of exposure to the toxic action of1Δg.Thus, our ESR and AFM results bring evidence that the multi-hydroxylated fullerene is an efficient1Δg-generator in aqueous media and might be implemented as a photosensitizer for performingoxidations in biological systems.