Spectral Characterization of Guanine C4-OH Adduct: A Radiation and Quantum Chemical Study

The reaction of hydroxyl radical ((OH)-O-center dot) with guanine was investigated under restricted pH condition (pH 4.6) using pulse radiolysis technique. The time-resolved optical transient absorption spectra showed two peaks centered at 300 and 330 nm at 4 mu s after the pulse which exhibited different reactivity toward molecular oxygen (O-2). The peak at 300 nm was found to be relatively more stable than the peak at 330 nm. The peak corresponding to 330 nm decayed within 20 mu s having a first order rate constant 4-7 x 10(4) s(-1) and was pH dependent. On longer time scale, the species decayed by a bimolecular process. The presence of O-2 did not affect its decay rate constant. The center dot OH reacts with guanine at pH 4.6 with a diffusion-controlled second order rate constant of >= 1 x 10(10) dm(3) s(-1). The reaction of Br-2(center dot-) O-2(center dot-) and 2-hydroxy-2-propyl radical with guanine was also investigated to differentiate among the one-electron oxidized, one-electron reduced species of guanine and the guanine OH adducts formed in the reaction of (OH)-O-center dot at pH 4.6. On the basis of the spectral characteristics and reactivity toward 02, two guanine-OH adduct species were identified (i) the C4-OH adduct species absorbing at 330 nm which has not been reported so far and (ii) the C8-OH adduct species absorbing at 300 rim in agreement with the known literature absorption features. Quantum chemical calculations using BHandHLYP with 6-31+G(d,p) basis set and excited state calculations using TDDFT for all possible transients complement the assignment of the observed spectral peak at 330 nm to the C4-OH adduct of guanine. Furthermore, steady state radiolysis revealed the formation of 8-hydroxy-guanine whose precursor is known to be the C8-OH adduct species.