DNA C1′ radicals lead to abasic site damage with the formation of 2-deoxyribonolactone residues. 1 Such alkaline-labile lesions result in strand scission 1 and have been reported to be mutagenic and resistant to repair nucleases. 2 The mechanistic aspects of C1′ radical reactions under either anoxic or aerobic conditions are currently under dispute. Figure 1 shows the reaction manifold for C1′ radicals illustrated for the specific case of the 2′-deoxyuridin-1′-yl radical (1). Reaction of 1 with a thiol such as glutathione returns the initial nucleoside or its anomer, whereas reaction with oxygen gives a C1′ peroxyl radical (3) that can ultimately lead to 2-deoxyribonolactone (6). These reactions have been discussed over the past two decades, 3 but quantitative kinetic measurements were not possible. Synthetic advances led to nucleosides modified with photoreactive groups that are specific C1′ radical precursors, and Greenberg 4,5 and Chatgilialoglu 6 reported product studies from radical 1, produced by photolysis of precursor 7, that are consistent with the general pathway in Figure 1. ESR and UV spectra of radical 1 were recently reported, and computational results revealed structural details of this radical. 7 In this work, we report the application of laser flash photolysis (LFP) methods for measurements of the kinetics of reactions of radical 1 with thiols and of superoxide release from peroxyl radical 3.Radical 1 was produced by 266-nm laser photolysis of precursor 7 as previously described. 7 The initial cleavage process must produce the pivaloyl radical, Me 3 CC(O)•, and 1 as the major products because no further growth in the UV spectrum of 1 was observed with ns-resolution after initial production by the laser flash ( Figure 2). The UV spectrum of radical 1 decays slowly in He-sparged solutions but faster in the presence of oxygen due to formation of peroxyl radical 3. A rate constant (k T ) of 1 × 10 9 M -1 s -1 was reported for the reaction of 1 with O 2 . 7 When radical 1 was produced in He-sparged solutions containing thiols, the rates of signal decay increased due to formation of 2 ( Figure 3A). Second-order rate constants for reactions of 1 at pH 7 and 20°C were (2.3 ( 0.5) × 10 6 M -1 s -1 for 2-mercaptoethanol, (2.9 ( 0.4) × 10 6 M -1 s -1 for cysteine, and (4.4 ( 0.3) × 10 6 M -1 s -1 for glutathione (errors at 2σ). The ratio of absolute rate constants for reaction of 1 with 2-mercaptoethanol and oxygen (k H /k T ) 2.3 × 10 -3 ) is in good agreement with the relative ratio found by Greenberg. 4 Rate constants for heterolytic fragmentation of peroxyl radical 3 (k f in Figure 1) to give superoxide radical anion, (O 2 ) •-, and cation 5 were determined from reactions conducted in the presence of tetranitromethane (TNM) which reacts with the superoxide radical anion to give the nitroform anion (eq 1) with λ max at 350 nm. 8,9 The TNM detection method is complicated. High concen- † Wayne State University. Gimisis, T.; Guerra, M.; Ferreri, C.; Emanuel, C. J.; Horner, J. H.; Newcomb, M.; Lucarin...

The thyminylmethyl radical and the 2′-deoxyuridin-1′-yl radical were studied. The former radical was produced in laser flash photolysis (LFP) studies from two precursors derived from thyminylacetic acid, the N-hydroxypyridine-2-thione ester (PTOC ester), and the phenylselenyl ester. The thyminylmethyl radical has an absorbance in the range 315-340 nm. The rate constant for its reaction with octadecanethiol in THF at ambient temperature determined by LFP methods is (3.1 ( 0.6) × 10 7 M -1 s -1 . The 2′-deoxyuridin-1′-yl radical was produced in bulk photolyses from both diastereomers of the corresponding C1′ tert-butyl ketone, 1′-pivaloyl-2′-deoxyuridine, and in LFP studies from one diastereomer. Trapping of this C1′ radical with 2-mercaptoethanol, cysteine, or glutathione gave both anomers of 2′-deoxyuridine; the product ratios were similar in each case and insensitive to precursor identity or thiol concentrations. Rate constants for reactions of the 2′-deoxyuridin-1′-yl radical with thiols and metal ions were determined by LFP methods; the respective rate constants for reactions with 2-mercaptoethanol, cysteine, glutathione, CuCl 2 , and FeCl 3 in water at ambient temperature are (2.3 ( 0.5) × 10 6 , (2.9 ( 0.4) × 10 6 , (4.4 ( 0.3) × 10 6 , (7.9 ( 0.3) × 10 7 , and ca. 1 × 10 8 M -1 s -1 . The 2′-deoxyuridin-1′-yl radical was addressed computationally. The radical center is not planar, and an energy profile for interconversion of the two anomeric forms of the radical was produced. Computed vertical transitions for the thyminylmethyl radical and one anomer of the 2′-deoxyuridin-1′-yl radical are in good agreement with the experimentally measured UV-visible spectra.

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