Amyloid-β-protein (Aβ) (25–35)-associated free radical generation is strongly influenced by the aggregational state of the peptides

We investigated whether or not the Amyloid-β-protein (Aβ) itself spontaneously generates free radicals using electron spin resonance (ESR) spectroscopy while also monitoring the aggregational state of Aβ and Aβ-induced cytotoxicity. The present results demonstrated a four-line spectrum in the presence of Aβ25–35 with N-tert-butyl-α-phenylnitrone (PBN) but not in the presence of PBN alone in phosphate-buffered saline (PBS). The fact that the four-line spectrum obtained for the Aβ25–35/PBN in PBS was completely abolished in the presence of the iron-chelating agent Desferal demonstrated the observed four-line spectrum to be iron-dependent. On the other hand, Aβ25–35 with PBN in phosphate buffer (PB) did not produce any definite four-line spectrum. the present results showed the amyloid fibril formation of Aβ25–35 in PBS to be much higher than that of Aβ25–35 in PB. Moreover, Aβ-induced cytotoxicity assays showed Aβ incubated in PBS to be more cytotoxic than that incubated in PB. These results thus demonstrate that Aβ(25–35) - associated free radical generation is strongly influenced by the aggregational state of the peptides.