SPECTROSCOPIC AND BIOLOGICAL TESTING OF PHOTOBLEACHING OF PORPHYRINS IN SOLUTIONS

The photobleaching of protoporphyrin IX (PP IX) and hematoporphyrin de rivative (HpD) solutions was followed using three different methods: s pectrophotometry, fluorometry and photodynamically induced cytotoxicit y. The latter entails photoirradiation of HT29 human colon adenocarcin oma cells in the presence of preirradiated solutions of HPD and PP IX (lambda less than or equal to 415 nm). The highest cytotoxicity was ob served in the presence of unirradiated dye and decreased with the time of preirradiation. This decay in photocytotoxicity was further used t o determine the porphyrin photobleaching kinetics in solution, For bot h sensitizers, quantum yields of photobleaching obtained by matching f luorescence were higher than that obtained from absorbance measurement s (10 and 11 times for HpD and PP IX, respectively), This difference r eflects preferential photobleaching of photolabile monomeric forms com pared to aggregates, The highest quantum yield was obtained in the bio logical test (decay in cytotoxicity) which was 14 times higher for HpD and 30 times higher for PP IX than the quantum yield obtained from ab sorbance measurements, The absence of correlation between biological a nd fluorescence measurements has to be taken into account in the in vi vo situation. Dark storage of preirradiated sensitizers (37 degrees C, 24 h) completely restored photocytotoxity for PP IX but only partiall y for HpD, whereas fluorescence patterns were partially restored for b oth sensitizers.