EPR and ENDOR study of radiation-induced radical formation in purines: sodium inosine crystals X-irradiated at 10 K.

X-irradiated single crystals of sodium inosine (Na(+)*Inosine(-)*2.5H(2)O), in which the hypoxanthine base is present as the N1-deprotonated anion, were investigated using K-band (24 GHz) electron paramagnetic resonance (EPR), electron-nuclear double resonance (ENDOR), and ENDOR induced EPR (EIE) techniques at 10 K. At least five different radicals were present immediately after irradiation at 10 K. R1, which decayed upon warming the crystals to 50 K, was identified as the electron-loss product of the parent N1-deprotonated hypoxanthine base. Hyperfine couplings to HC8 and HC2 were fully characterized with ENDOR spectroscopy, and the identification was supported by DFT calculations. R2, which also decayed on warming to 50 K, exhibited nearly equal couplings to HC2 and HC8. Taken in combination with an extensive set of DFT calculations, the experimental results indicate that R2 is the (doubly negative) product of electron-gain by the initially anionic N1-deprotonated hypoxanthine parent. R3, which exhibited hyperfine coupling only to HC8 could not be identified. R4, which persisted on annealing to 260 K, exhibited one large alpha-proton hyperfine coupling which was fully characterized by ENDOR. Based on DFT calculations and the experimental data, R4 was identified as the product of net H-abstraction from C5'. The remaining HC5' was the source of the measured alpha-proton coupling. R5, present at low temperature and the only observable radical after warming the crystals to room temperature, was identified as the C8-H addition radical. The alpha-coupling to HC2 and beta-couplings to the pair of C8 methlyene protons were fully characterized by ENDOR.

Radiation-induced hydroxyl addition to purine molecules: EPR and ENDOR study of hypoxanthine hydrochloride monohydrate single crystals.

Three radical species were detected in an EPR/ENDOR study of X-irradiated hypoxanthine.HCl.H2O single crystals at room temperature: RI was identified as the product of net H addition to C8, RII was identified as the product of net H addition to C2, and RIII was identified as the product of OH addition to C8. The observed set of radicals was the same for room-temperature irradiation as for irradiation at 10 K followed by warming the crystals to room temperature; however, the C2 H-addition and C8 OH-addition radicals were not detectable after storage of the crystals for about 2 months at room temperature. Use of selectively deuterated crystals permitted unique assignment of the observed hyperfine couplings, and results of density functional theory calculations on each of the radical structures were consistent with the experimental results. Comparison of these experimental results with others from previous crystal-based systems and model system computations provides insight into the mechanisms by which the biologically important purine C8 hydroxyl addition products are formed. The evidence from solid systems supports the mechanism of net water addition to one-electron oxidized purine bases and demonstrates the importance of a facial approach between the reactants.

EPR and ENDOR study of radiation-induced radical formation in purines: hypoxanthine hydrochloride monohydrate crystals X-irradiated at 10 K.

Electron paramagnetic resonance (EPR) and electron-nuclear double resonance (ENDOR) study of hypoxanthine.HCl.H(2)O crystals irradiated at low temperatures (10 K) identified three radical species. In these crystals, the parent molecules exist in a cationic form with a proton at N7. R1 was the product of net hydrogen addition to N3 and exhibited alpha-proton hyperfine couplings to HC2, HN1, HC8, and HN3. The coupling to HC2 has an isotropic component smaller than usual, evidently an indication that the bonds to C2 are nonplanar. R2 was the product of net hydrogen loss from N7, equivalent to the one-electron oxidation product of neutral hypoxanthine, and exhibited alpha-proton hyperfine couplings to HC2 and HC8. Both couplings are characteristic of planar bonding arrangements at the centers of spin. R3 was provisionally identified as the product of net hydrogen addition to O6 and exhibited hyperfine alpha-proton couplings to HC8 and NH1. To identify the set of radicals, the experiments employed four crystal types: normal, deuterated only at NH positions, deuterated at HC8 and NH positions, and deuterated at HC8 only. The low-temperature data also showed clear evidence for H/D isotope effects in formation and/or stabilization of all radicals. To aid and support the identifications, the experimental results were compared to DFT calculations performed on a variety of radical structures plausible for the parent molecule and molecular packing within the crystal.

Electronic g-factor measurement from ENDOR-induced EPR patterns: malonic acid and guanine hydrochloride dihydrate.

Measurement of electronic g-factors (g) from radicals in irradiated organic crystals is generally difficult because the overall EPR pattern is usually the composite of several components, e.g., from multiple radicals and from multiple magnetic sites. However, when an ENDOR line is fully resolved, the method of ENDOR-induced EPR (EI-EPR, or EIE) in principle permits identification of the EPR pattern from the individual component yielding the line. To examine this method as an approach useful for measuring g, we used it to measure those of known radicals in two different crystal systems. First, to verify correspondence of the EIE and EPR sufficient for using EIE patterns to extract g, we used both EIE and EPR to measure g of (*CH(COOH)(2) from irradiated crystals of malonic acid. Then, to illustrate the procedure applied to a system giving a more complex EPR pattern, we used EIE to measure g of the O6-protonated anion radical of guanine in irradiated guanine.HCl.2H(2)O crystals. EPR results from the malonic acid radical are g(max)=2.00374(2), g(mid)=2.00331(2), and g(min)=2.00234(3); EIE results from the same radical are g(max)=2.00375(2), g(mid)=2.00334(2), and g(min)=2.00238(2), where numbers in parentheses indicate statistical uncertainties in the respective least significant digits. In addition, eigenvectors from the two sets of measurements agree to approximately 1 degrees. Results from the guanine radical are g(max)=2.00490(2), g(mid)=2.00318(4), and g(min)=2.00218(4). (The uncertainties should reliably indicate relative accuracy, while absolute accuracy is within +/-0.0002 as indicated by simultaneous measurement of Cr(3+) in MgO.)