Phototoxicity mechanism of a kryptocyanine dye in human red cell membranes and isolated murine mitochondria.

The phototoxicity mechanism of a kryptocyanine dye, N,N'-bis(2-ethyl-1,3-dioxolane)kryptocyanine (EDKC+), has been studied in RBC membranes and isolated mitochondria. Lipophilic, positively charged dyes, such as EDKC+, may be useful as tumor-cell-selective, light-activated cytotoxic agents. Exposure of the RBC membranes to 700-nm light and EDKC+ inhibited membrane acetylcholinesterase and photodecomposed EDKC+ in air-purged but not argon-purged samples. Photoinactivation of acetylcholinesterase was the same in D2O as in H2O and was not quenched by superoxide dismutase. Ascorbate and azide (10 mM) quenched or slightly enhanced, respectively, the inactivation. In argon-purged samples containing methyl viologen, EDKC+ photodecomposed, but acetylcholinesterase activity was unaffected. The mechanism may involve electron transfer to oxygen and subsequent formation of toxic photoproducts from EDKC+. In contrast, exposure of murine mitochondria to EDKC+ and 700-nm light caused inhibition of mitochondrial respiration in both the presence and absence of oxygen. The photodecomposition of EDKC+ correlated with inhibition of respiration. Thus, the phototoxicity of EDKC+ in mitochondria may be due to electron transfer from photoexcited EDKC+ to oxygen and electron acceptors in the membrane. These studies indicate that dyes such as EDKC+ may be useful for photochemotherapy of hypoxic regions in tumors.

Pharmacological studies of arrhythmias induced by rose bengal photoactivation.

Singlet oxygen and superoxide production by rose bengal photoactivation leads to rapid electrophysiological changes and arrhythmias. To investigate which intermediate is causative and to probe possible mechanisms, hearts (n = at least 6/group) were perfused aerobically for 10 min without rose bengal followed by 5 min with rose bengal before illumination for 20 min. In controls, all or most hearts exhibited ventricular premature beats, ventricular tachycardia, and complete atrioventricular block. Most antioxidants tested had no protective effect; histidine, however, significantly delayed the onset of electrocardiographic (ECG) changes. In further studies, two antiarrhythmic agents (quinidine and verapamil) had no little protective effect, whereas R56865 significantly delayed the onset of ECG changes and reduced the incidence of arrhythmias. However, spectrophotometric and laser pulse radiolysis studies showed that this apparent protective effect might have resulted from an interaction between R56865 and the rose bengal molecule, leading to a reduction in singlet oxygen production. In conclusion, the electrophysiological changes induced by rose bengal photoactivation are likely to be due to singlet oxygen; antiarrhythmic drugs appear to be unable to protect against the injury unless there is some interaction with the photoactivation process.

Partition of rose bengal anion from aqueous medium into a lipophilic environment in the cell envelope of Salmonella typhimurium: implications for cell-type targeting in photodynamic therapy.

Photodynamic therapy employs photosensitizers for the selective destruction of tumor tissue while sparing the surrounding healthy tissue. Photosensitization may also be applied to the selective eradication of microorganisms. Photosensitized inactivation requires that the sensitizer bind to the target and therefore the factors that determine photosensitizer binding are critical to photosensitization selectivity. This paper reports the determination of some features of the binding site of the potent photosensitizer, Rose Bengal, in Salmonella bacteria and describes some of the factors that affect this binding. The shift in the wavelength of maximum fluorescence and experiments with the fluorescence quencher TNBS indicate that Rose Bengal is located in a non-aqueous compartment such as the outer membrane. The dye does not seem to significantly accumulate inside the cell, but rather to accumulate in the outer membrane. Time-dependent changes in sensitizer localization in two strains of Salmonella typhimurium that differ in cell wall formation, LT-2 and TA1975, correspond to their differences in susceptibility to photosensitized killing. Therefore these results provide clues to the factors that determine photosensitization selectivity. Understanding this phenomenon is essential for the efficient design of selective photosensitizers and for optimizing antitumor and antiviral photodynamic therapy.