Abstract—In this work we have produced the π‐cation radicals of a number of nucleotides, dinucleoside phosphates, and DNA in aqueous glasses (8M NaCIO4) by photoionization and investigated these species by EPR spectroscopy. Results found for nucleotides and dinucleoside phosphates containing one type of DNA base, e.g. TpT, GpG, or dApdA, were used in the analysis of spectra found for mixed dinucleoside phosphates, e.g. TpdG. For TpdG and TpdA in neutral glasses photoionization takes place from the purine base and no transfer of charge to the pyrimidine base is found. In basic conditions both the adenine and thymine π‐cations are observed in TpdA. In both neutral and basic conditions the results found for mixed dinucleoside phosphates containing guanine show that the guanine cation is formed preferentially by photolysis. This result was found to extend to DNA. Photolysis of DNA in8MNaC1O4produced principally the guanine cation. Computer simulations using parameters determined by other workers from a study of γ‐irradiated oriented DNA closely match the spectrum found in this work attributed to the guanine cation in dinucleoside phosphates and DNA. This work thus confirms the presence of the guanine cation in γ‐i