Cultured human lymphoblasts (CCRF-CEM line) have DNA glycosylase activities, the specificities of which were investigated by using high-performance liquid chromatography. In addition to 3-methyladenine, 3-ethyladenine, 7-methylguanine, 7-ethylguanine, 3-methylguanine, and 3-ethylguanine also were excised from alkylated double-stranded DNA and deoxypolynucleotides, but 1-methyladenine, 1-ethyladenine, O6-methylguanine, and O6-ethylguanine were not. The glycosylase activity was generally greater for the methylated than for the corresponding ethylated purines and was also greater toward 3-alkyladenine than toward 3-alkyladenine than toward 3-alkylguanine. 7-Methylguanine and 7-ethylguanine were excised to similar but low extents. However, in molar terms, the release of 7-methylguanine was similar to that of 3-methyladenine.