Abstract
Identification of the radicals was performed for the standard reaction mixtures, which contained 4.3 mM oleic acid, 25 µM riboflavin, 1 mM FeSO(4)(NH(4))(2)SO(4), 10 mM cholic acid, 40 mM phosphate buffer (pH 7.4) and 0.1 M α-(4-pyridyl-1-oxide)-N-tert-butylnitrone under the UVA irradiation (365 nm), using an electron spin resonance, an high performance liquid chromatography-electron spin resonance and an high performance liquid chromatography-electron spin resonance-mass spectrometry. The electron spin resonance and high performance liquid chromatography-electron spin resonance measurements of the standard reaction mixtures showed prominent signals (α(N) = 1.58 mT and α(H)β = 0.26 mT) and peaks 1 and 3 (retention times, 37.0 min and 49.0 min). Since the peak 3 was not observed for the standard reaction mixture without oleic acid, the radical of the peak 3 seems to be derived from oleic acid. Singlet oxygens seem to participate in the formation of the oleic acid-derived radicals because the peak height of the peak 3 observed in the standard reaction mixture of D(2)O increased to 308 ± 72% of the control. The high performance liquid chromatography-electron spin resonance-mass spectrometry analysis showed that 7-carboxyheptyl radical forms in the standard reaction mixture.