Pyrimidines radical cations

A different strategy has been applied in our work, that emphasizes the importance of DNA stability on hole transfer within double-stranded DNA. This work is based on determination of the overall yield of oxidized nucleosides that arise from the conversion of initially generated purine and pyrimidine radical cations within DNA exposed to two-photon UVC laser pulses. On the one hand, this work benefits from the excellent current knowledge of chemical reactions involving the radical cations of DNA bases, and on the other hand, from major analytical improvements that include recent availability of the powerful technique of high performance liquid chromatography-electrospray ionization-tandem mass spectrometry (CLHP-ESI-MS/MS) [16-18]. 

A number of papers have reported studies on pyrimidine radical cations. 1-Methylthymine radical cations generated via a triplet-sensitized electron transfer to anthraquinone-2,6-disulfonic acid were detected by Fourier transform electron paramagnetic resonance (FTEPR). The parent 1-methylthymine radical cation, and its transformation to the N(3)-deprotonated radical cation, were observed. Radical cations formed by addition of HO and POs" at C(6) were also detected depending on the pH. Similarly, pyrimidine radical cations deprotonated at N(l) and C(5)-OH were detected from the reaction of 804 with various methylated pyrimidines." These radicals are derived from the initial SO4 adducts of the pyrimidines. Radical cations of methylated uracils and thymines, generated by electron transfer to parent ions of... 

Short-lived adducts may be formed as intermediates in the reactions of the oxidizing inorganic radicals with the nucleobases, and it is therefore not always fully excluded that processes observed at very short times and attributed to the reactions of radical cations are in fact due to such intermediates. It may be mentioned that, for example, a long-lived S04 -adduct is observed in the reaction of S04, with maleic acid (Norman et al. 1970). It has been suggested that S04, in its reactions with the pyrimidines forms only an adduct and does not give rise to radical cations (Lomoth et al. 1999). The observation of heteroatom-centered radicals by EPR from the nucleobases Ura, Thy and Cyt (Catterall et al. 1992) as well as dCyd (Hildenbrand et al. 1989) (see below) has been taken as evidence that in the reaction of S04 with pyrimidines radical cations are likely, albeit... 

Pyrimidines. Photoexcited anthraquinone-2,6-disulfonate undergoes ET with Thy and its methyl derivatives as indicated by Fourier transform EPR (Geimer et al. 1997). These pyrimidine radical cations deprotonate at N( 1) thereby giving rise to the corresponding N-centered radicals [reaction (6)]. 

It has been mentioned above that the pyrimidine radical cations are reasonably strong acids and rapidly deprotonate at a heteroatom. As all protonation/ deprotonation reactions at heteroatoms are reversible [e.g., equilibrium (22)], the radical cations are regenerated upon reprotonation. Deprotonation at carbon or reaction with water yield the final free-radical products. For the l,3Me2Thy system, where the deprotonation/reprotonation equilibria such as reaction (22) fall away, reactions (25)-(28) have been postulated to account for the fact that in the presence of 02 l,3Me25HOMeUra and l,3Me25(CHO)Ura [reaction (29)] are formed in a combined yield of 80% of primary S04 radicals (Rashid et al. 1991). The formation of these products has been taken as evidence that a free radical cation must be an intermediate. It is, however, also possible that the allylic radical is formed in a concerted reaction HS04 elimination. For such a process, a six-membered transition state can be written. 

The radical anions may be formed by reacting the nucleobases with eaq which may be either generated radiolytically or in a two-step reaction, e.g in the laser flash photolysis of anthraquinonedisulfonate in the presence of pyrimidines (yielding the pyrimidine radical cation and an anthraquinonedisulfonate radical anion) and subsequent photoionization of the anthraquinonedisulfonate radical anion (Lii et al. 2001). The latter approach, combined with Fourier transform EPR spectroscopy, yielded detailed information as to the conformation of the radical anions of Ura and Thy in aqueous solution (for a discussion see Close 2002 Naumov and Beckert 2002). Similarly valuable EPR information has been obtained from y-irradiated single crystals (cf. Box and Budzinski 1975 Boxet al. 1975 Sagstuen et al. 1998). 

Decarroz C, Wagner JR, Cadet J (1987) Specific deprotonation reactions of the pyrimidine radical cation resulting from the menadione mediated photosensitization of 2 -deoxycytidine. Free Rad Res Commun 2 295-301... 

Hydration of the thymidine radical cation leads to the predominant formation of the oxidizing 6-hydroxy-5,6-dihydrothymid-5-yl radical whereas deprotonation mostly generates the methyl-centered radical (Fig. 4). As already pointed out, these two pyrimidine radicals are also produced upon reaction with OH. However, the methyl-centered radical represented only 5% of base radicals when produced by OH while it corresponds to 30% of the thymidine radicals produced upon one-electron oxidation. Type I photosensitization of 2 -deoxy-cytidine leads to the formation of the 6-hydroxy-5,6-dihydro-2 -deoxycytidil-5-yl radical also produced upon reaction with OH. Additional oxidative pathways involve the formation of 2-deoxyribonolactone and free cytosine as well as production of 2 -deoxyuridine as the result of deprotonation ofthe pyrimidine radical cation at Cl and NH2 group, respectively. 

Pulse radiolysis shows that the pyrimidine radical cations are fairly strong acids and rapidly deprotonate at a heteroatom [reaction (98)]. As protonation/deprotonation reactions at heteroatoms are easily reversible, the radical cations are regenerated upon reprotonation. Deprotonation at carbon or reaction with water yields the final free-radical products [reactions (99) - (101)]. It is noted that in thymidine [23] and 5 -thymidylic acid [104] the allylic thymine radical is observed by EPR and there is very little question that its precursor is the thymine radical cation. The identification of the C(6)-OH-5-yl radical by EPR supports the view [100] that reaction with water competes with the deprotonation at methyl. Due to the ready oxidation of the (reducing) C(5)-OH-6-yl by peroxodisulfate, this type of radical is only observed at low peroxodisulfate concentrations in these systems, i.e. the (oxidizing) C(6)-OH-5-yl radicals are correspondingly enriched under conditions favourable to a chain reaction [22]. In the case of 1,3-dimethyluracil the interesting characteristics of... 

Scheme 2 Pyrimidine radical cation formation and reactivity.

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