The UVA and UVB components of sunlight can produce three classes of bipyrimidine DNA photolesions [cyclobutane pyrimidine dimers (CPDs), pyrimidine (6-4) pyrimidone photoproducts (6-4PPs) and related Dewar valence isomers (DewarPPs)]. The UVA/UVB ratio of sunlight is high in winter and low in summer in the Northern Hemisphere. Since UVB radiation produces 6-4PPs and UVA radiation converts them into DewarPPs through photoisomerization, it is expected that there may be differences in the photoisomerization of 6-4PPs between summer and winter, although that has never been documented. To determine that, isolated DNA was exposed to natural sunlight for 8 h in late summer and in winter, and absolute levels of the three classes of photolesions were quantified using calibrated ELISAs. It was found that sunlight produces CPDs and 6-4PPs in DNA at a ratio of about 9:1 and converts approximately 80% of 6-4PPs into DewarPPs within 3 h. Moreover, photoisomerization is more efficient in winter than in late summer after sunlight irradiation for the same duration, at similar solar UV doses and with the same induction level of CPDs. These results demonstrate that seasonal differences in the UVA/UVB ratio influence the efficiency of the photoisomerization of 6-4PPs into DewarPPs.