This study determined whether nascent chromatin in nuclei from leukemia L1210 cells constitutes a preferential target for bleomycin. No differences were seen in fragmentation of nascent and bulk DNA as judged by DNA double-stranded cleavage and the release of acid-soluble material or subnucleosomal (under 8 S) fragments. In contrast, bleomycin-induced chromatin aggregation (Woynarowski, J.M., Gawron, L.S. and Beerman, T.A. (1987) Biochim. Biophys. Acta 910, 149-156) occurred preferentially in nascent chromatin as indicated by a retarded solubilization of nascent chromatin and generation of a fast-sedimenting material (above 45 S) in the sedimentation profiles of drug-released nascent chromatin. This preferential aggregation disappeared completely when chromatin became older than 10 min. The drug aggregation activity did not distinguish nascent and mature presolubilized oligonucleosomes. The results suggest that bleomycin recognizes higher-order structures of nascent chromatin.