Characterization of the high-resolution ESR spectra of superoxide radical adducts of 5-(diethoxyphosphoryl)-5-methyl-1-pyrroline N-oxide (DEPMPO) and 5,5-dimethyl-1-pyrroline N-oxide (DMPO).: Analysis of conformational exchange

It has been previously reported that the spin trap 5-(diethoxyphosphoryl)-5-methyl-1-pyrroline N-oxide (DEPMPO) can form stable radical adducts with superoxide radical. However, the presence of diastereomers of DEPMPO radical adducts and the appearance of superhyperfine structure complicates the interpretation of the ESR spectra. It has been suggested that the superhyperfine structure in the ESR spectrum of DEPMPO/(OOH)-O-center dot is a result of conformational exchange between conformers. The analysis of the temperature dependence of the ESR spectrum of DEPMPO/(OOH)-O-center dot and of its structural analog DMPO/(OOH)-O-center dot have demonstrated that both ESR spectra contain exchange effects resulting from conversion between two conformers. Computer simulation calculates a conformer lifetime on the order of 0.1 mu s for DMPO/(OOH)-O-center dot at room temperature. However, temperature dependence of the ESR spectrum of DEPMPO/(OOH)-O-center dot suggests that superhyperfine structure does not depend on the conformational exchange. We have now found that the six-line ESR spectrum with superhyperfine structure should be assigned to a DEPMPO-superoxide-derived decomposition product. Therefore, ESR spectra previously assigned to DEPMPO/(OOH)-O-center dot contain not only the two diastereomers of DEPMPO/(OOH)-O-center dot but also the decomposition product, and these spectra should be simulated as a combination of four species: two conformers of the first diastereomer, one conformer of the second diastereomer and the superoxide-derived decomposition product. The presence of four species has been supported by the temperature dependence of the ESR spectra, nucleophilic synthesis of radical adducts, and isotopic substitution experiments. It is clear that to correctly interpret DEPMPO spin trapping of superoxide radicals, one must carefully consider formation of secondary radical adducts.