Binding studies of porphyrins to human serum albumin using affinity capillary electrophoresis

The present work demonstrates that affinity capillary electrophoresis (ACE) can be employed as a valuable and powerful tool for studying the interacti ons between porphyrins and proteins in biological and biomedical research, such as the development of porphyrins and related compounds as efficient an d selective photosensitizers in the photodynamic therapy of cancers. Bindin g constants of human serum albumin (HSA) to four biological porphyrins (uro porphyrin 1, heptacarboxylporphyrin, coproporphyrin I, protoporphyrin IX), which possess a wide range of hydrophobicity, were estimated by ACE. Based on 1:1 molecular association between these individual porphyrins and HSA, t he change of the electrophoretic mobility of HSA as a function of porphyrin concentration in the run buffer was measured and the binding constants wer e calculated from the slope of the Scatchard plots. The binding constant va lues were found to be 8.80 +/- 0.51 x 10(4) m(-1), 2.39 +/- 0.16 x 10(5) m( -1), 1.61 +/- 0.11 x 10(6) m(-1), and 9.34 +/- 0.30 x 10(6) m(-1) for uropo rphyrin I, heptacarboxylporphyrin, coproporphyrin I, and protoporphyrin IX, respectively, and most of these results are in good agreement with those r eported in the literature using conventional methods for binding measuremen ts. Additionally, experimental binding constant data obtained using ACE was found to exhibit very good correlation with theoretical hydrophobicity val ues calculated using the Rekker's hydrophobic fragmental constant method, t hus further supporting the hypothesis that the hydrophobicity of the porphy rin side chains play an important role in governing the hydrophobic interac tion of porphyrins with serum proteins such as HSA.