ESR detection of OH radicals generated inside cells by X-ray and evaluation of the cytotoxicity of DMPO

Abstract

An attempt was made to identify OH radicals inside cells in which the radicals were generated by X-ray radiolysis. In the experiments, HL60 cells were incubated for 1 hour in a medium containing 100mM 5,5-dimethyl-a-pyrroline-1-oxide (DMPO). Cells were subjected with 20Gy radiation exposure after which the cells were centrifuged. The centrifuged cells were packed in an ESR quartz column and the ESR spectrum was measured. The ESR spectrum of OH radicals was obtained 5 minutes after the irradiation; however, the ESR spectrum of DMPO-OH was weak and decreased rapidly with time in this cell-containing system. The results indicate that the main difficulty in applying the spin-trapping technique to oxygen-derived radicals formed in cell arise from the chemical instability of the resulting spin adducts. To improve the technique to study cellular systems, it will be necessary to increase the stability of the generated spin adduct.