Hydogen peroxide-dependent photocytotoxicity by phloxine B, a xanthene-type food colorant

Background: Phloxine B (PhB; 2′,4′,5′,7′- tetrabromo-4,5,6,7-tetrachloro-fluorescein), an artificial xanthene colorant, has been used as a red coloring agent in drugs and cosmetics as well as foods in some countries. However, little effort has been devoted to the study of this colorant as a potentially useful medicinal agent. Methods: We investigated the daily light-induced photocytotoxicity of PhB in two human leukemia cells, HL-60 and Jurkat, and its underlying mechanisms by in vitro experiments using antioxidants. Reuslts and conclusions: PhB inhibited cell proliferation more preferentially to HL-60 cells than to Jurkat cells. Co-treatment of catalase completely blocked the photocytotoxicity by PhB in HL-60 cells, whereas the effect of histidine was only partial, suggesting that hydrogen peroxide (H ₂O₂), rather than singlet oxygen, might be a prerequisite for the PhB-induced HL-60 cell death. Actually, PhB produced a significant amount of H₂O₂ in the media as well as in the cells in concentration- and light-dependent manners. Furthermore, methionine, a hypochlorous acid (HOCl) scavenger, also significantly attenuated the cytotoxicity in HL-60 cells, but not in Jurkat cells, indicating the involvement of myeloperoxidase (MPO)-dependent hypohalous acid formation during the photocytotoxicity. In vitro experiments revealed that halogenated tyrosine was generated from the reaction of bovine serum albumin with PhB and HL-60 cell lysate. The present findings suggested that PhB induced a differential photodynamic action in the MPO-containing leukemia cells through an H ₂O₂-dependent mechanism. General significance: Our findings provide new insights into the molecular mechanisms underlying the PhB-induced apoptosis and also evaluated PhB as a promising PDT agent.