pH-Dependent Singlet O 2 Oxidation Kinetics of Guanine and 9‑Methylguanine: An Online Mass Spectrometry and Spectroscopy Study Combined with Theoretical Exploration Wenchao Lu, †,‡ Yan Sun, †,‡ Wenjing Zhou, † and Jianbo Liu* ,†,‡ † Department of Chemistry and Biochemistry, Queens College of the City University of New York, 65-30 Kissena Boulevard, Queens, New York 11367, United States ‡ Ph.D. Program in Chemistry, The Graduate Center of the City University of New York, 365 Fifth Avenue, New York, New York 10016, United States * S Supporting Information ABSTRACT: We report a kinetic and mechanistic study on the title reactions, in which 1 O 2 was generated by the reaction of H 2 O 2 with Cl 2 and bubbled into an aqueous solution of guanine and 9-methylguanine (9MG) at different pH values. Oxidation kinetics and product branching ratios were measured using online electrospray ionization mass spectrometry coupled with absorp- tion and emission spectrophotometry, and product structures were determined by collision-induced dissociation (CID) tandem mass spectrometry. Experiments revealed strong pH dependence of the reactions. The oxidation of guanine is noticeable only in basic solution, while the oxidation of 9MG is weak in acidic solution, increases in neutral solution, and becomes intensive in basic solution. 5-Guanidinohydantoin (Gh) and spiroiminodihy- dantoin (Sp) were detected as the major oxidation products of guanine and 9MG, and Sp became dominant in basic solution. A reaction intermediate was captured in mass spectra, and assigned to gem-diol on the basis of CID measurements. This intermediate served as the precursor for the formation of Gh. After taking into account solution compositions at each pH, first-order oxidation rate constants were extracted for individual species: that is, 3.2-3.6 × 10 7 M -1 s -1 for deprotonated guanine, and 1.2 × 10 6 and 4.6-4.9 × 10 7 M -1 s -1 for neutral and deprotonated 9MG, respectively. Guided by approximately spin- projected density-functional-theory-calculated reaction potential energy surfaces, the kinetics for the initial 1 O 2 addition to guanine and 9MG was evaluated using transition state theory (TST). The comparison between TST modeling and experiment confirms that 1 O 2 addition is rate-limiting for oxidation, which forms endoperoxide and peroxide intermediates as determined in previous measurements of the same systems in the gas phase. 1. INTRODUCTION Oxidatively generated damage of DNA nucleobases gives rise to mutagenesis, DNA-protein cross-linking, and cellular lethality. 1-3 One category of such damage is caused by electronically excited singlet oxygen (O 2 ,a 1 Δ g ) which is produced by photosensitization and/or a range of enzymatic and nonenzymatic reactions in live organisms. 4 Of the four DNA nucleobases (i.e., adenine, thymine, guanine, and cytosine), guanine (G) is the most susceptible to 1 O 2 oxidation. The oxidation mechanism of guanine nucleoside is outlined in Scheme 1. 5-17 In brief, deoxyguanosine (dGuo) is attacked by 1 O 2 on the imidazole ring for a [4 + 2] cycloaddition, forming a transient endoperoxide that quickly converts to a hydroperoxide 8-OOHdGuo. 8-OOHdGuo within DNA is mainly reduced to 8-oxo-7,8-dihydrodeox- yguanosine (OdGuo). Free 8-OOHdGuo or that in short oligonucleotides, on the other hand, undergoes dehydra- tion to form oxidized 8-oxo-7,8-dihydrodeoxyguanosine (OdGuo ox ) and then rehydration to form 5-hydroxy-8-oxo-7,8- dihydrodeoxyguanosine (5-OHOdGuo). Subsequent conver- sions from 5-OHOdGuo are pH-dependent. Under basic conditions, 5-OHOdGuo goes through an acyl shift to produce spiroiminodihydantoin (dSp); 18 whereas under acidic conditions, the formation of a gem-diol intermediate via the addition of a water to 5-OHOdGuo becomes predominant. Ring-opening at the N1-C6 bond of gem-diol, accompanied by an intramolecular proton transfer, leads to a 4-carboxydGh. Decarboxylation of the latter results in the formation of 5-guanidinohydantoin (dGh). 9,19 In addition, OdGuo may react with a second 1 O 2 and form 5-hydroperoxy-8-oxo-7,8-dihydrodeoxyguanosine (5-OOHOdGuo), followed by reduction to 5-OHOdGuo or decarboxylation to oxidized 5-guanidinohydantoin (dGh ox ). 14 Received: September 25, 2017 Revised: November 28, 2017 Published: November 29, 2017 Article pubs.acs.org/JPCB Cite This: J. Phys. Chem. B 2018, 122, 40-53 © 2017 American Chemical Society 40 DOI: 10.1021/acs.jpcb.7b09515 J. Phys. Chem. B 2018, 122, 40-53