Deprotonation of the adenine (A) base in both mononucleotide and oligonucleotide (ODN) was measured by nanosecond pulse radiolysis. The cation radical (A(+center dot)) of deoxyadenosine (dA), produced by oxidation with SO4-center dot, rapidly deprotonated to form the neutral A radical (A(-H)(center dot)) with a rate constant of 2.0 x 10(7) s(-1) and a pK(a) value of 4.2, as determined by transient spectroscopy. A similar process was observed in experiments performed on a variety of double-stranded ODNs containing adenine center dot thymine (A center dot T) base pairs. The transient spectrum of A(+center dot) in an ODN composed of alternating A center dot T pairs was essentially identical to that of free dA and differed from the spectra of ODNs containing AA and AAA. In contrast, the spectra of A(-H)(center dot) were not affected by the sequence. These results suggest that the positive charge on A(+center dot) in ODNs is delocalized as the dimer is stabilized by pi-orbital stacking between adjacent A's. The rate constants for deprotonation of A(+center dot) in ODNs containing AA and AAA (0.9-1.1 x 10(7) s(-1)) were a factor of 2 smaller than the rate constants for deprotonation of A(+center dot) in ODNs containing alternating A center dot T and dA (2.0 x 10(7) s(-1)). This suggests that the formation of a charge resonance stabilized dimer AA(+center dot) in DNA produced a significant barrier to deprotonation.