A hematoporphyrin derivative (abbreviated in the literature as "HPD") has been used successfully for phototherapy of tumors in the clinic. The chemical nature of HPD, a complex mixture of porphyrins, is not fully understood. This study was designed to provide an explanation for the superior tumor-localizing ability of HPD. Chromatographic behavior, hydrophobicity, transport, binding, and photosensitizing capacity of different porphyrins were examined and compared. Biological studies were carried out using murine leukemia L1210 cells in vitro. The initial rate of porphyrin uptake was a function of drug hydrophobicity. The most hydrophobic components of HPD were therefore the most potent photosensitizers when irradiation followed a 10-min porphyrin-loading incubation. But these and other hydrophobic porphyrins were readily washed from cells by medium containing serum. Hydrophilic components of HPD were gradually accumulated by L1210 cells via a mode of binding not readily dissociated by washing and appear to be responsible for the preferential affinity of this product for neoplastic cells. A portion of the tightly bound porphyrin could not be dissociated by sodium dodecyl sulfate:polyacrylamide gel electrophoresis but remained bound to a low-molecular weight membrane component.