STUDIES ON THYMINE AND URACIL TRIPLET EXCITED STATE IN ACETONITRILE AND WATER

Abstract— Thymine and uracil triplet-triplet absorption spectra and triplet excited state lifetimes have been observed in acetonitrile and water by nanosecond laser flash spectroscopy. A study of triplet energy transfer from these pyrimidines to retino! has also allowed an estimation of the triplet extinction coefficient ε_TT of thymine and uracil. These ε_TT were then used to determine the triplet quantum yields Ø_T of both pyrimidines in acetonitrile and water.     

THE KINETICS OF REDUCTION OF CYTOCHROME c BY SEMI-REDUCED FLAVIN

Abstract— Flavin mononucleotide radicals, FMNH', generated by laser flash photolysis of FMN in the presence of the electron donors, histidine, guanosine monophosphate or EDTA, were found to reduce cytochrome c with an apparent rate constant of 6 ± 10⁷ M ⁻¹ s⁻¹. These flash photolysis results were, however, complicated by the electron donor radicals formed simultaneously which, particularly with EDTA, also lead to reduction of cytochrome c. Pulse radiolysis of a nitrous oxide saturated aqueous solution of FMN containing a high concentration of HCOONa, leads to the exclusive formation of FMNH'. By adding small concentrations of cytochrome c to this solution, a rate constant of 4.0 ± 10⁻¹ M ⁻¹ s⁻¹ was obtained for the reduction of cytochrome c by FMNH'. Replacement of the HCOONa by EDTA in such solutions leads to further routes for reduction of cytochrome c on radiolysis. as in the photolytic situation. The relevance of these results to flavin-photosensitised reduction of cytochrome c and other components of the mitochondrial electron transport chain is discussed.     

ANOXIC PHOTODAMAGE IN THE PRESENCE OF PORPHYRINS: EVIDENCE FOR THE LACK OF EFFECTS ON MITOCHONDRIAL MEMBRANES

Abstract When isolated mitochondria are irradiated in the presence of 50 μg/ml hematoporphyrin or hematoporphyrin derivative in the respiratory medium, the irradiation is at least twenty-fold less effective in impairing the Ca²⁺ pump in the absence of oxygen than in its presence. This result suggests that photosensitization by non oxygen-depending pathways has few or no effects on cell membranes, a result somewhat at variance with those observed with liposomes.     

Ueber die Spectra des Schwefels, Selens und Tellurs

Ohne Zusammenfassung     

Carrier thiols are targets of Photofrin II photosensitization of isolated rat liver mitochondria

To gain further insight into the ability of Photofrin II to photosensitize mitochondrial translocators, and to ascertain whether mitochondrial thiols are specific targets of Photofrin II, the activity of phosphate carrier was measured in isolated rat liver mitochondria irradiated with 365 nm light in the presence of Photofrin II. Photodynamic treatment decreased the maximum rate of phosphate uptake, without changing the phosphate affinity for its own carrier. The ability of the thiol reagent mersalyl (an inhibitor of phosphate, dicarboxylate and oxodicarboxylate carriers) to protect these carriers against Photofrin II photosensitization was also tested. Protection was observed, indicating the involvement of carrier thiols in mitochondrial photosensitization.     

Zur spectroskopischen Nachweisung des Kupfers

Ohne Zusammenfassung     

PHOTODYNAMIC EFFECTS INDUCED BY FUROCOUMARINS ON A MEMBRANE SYSTEM. COMPARISON WITH HEMATOPORPHYRIN

Abstract Isolated rat liver mitochondria have been used as a model to investigate the photodynamic action of psoralen (Ps) and 4,5',8-trimethylpsoralen (TMP) on cellular membrane systems in comparison with hematoporphyrin (Hp). Oxidation was detected by the consumption of free oxygen as measured polarographically in the respiratory chamber when irradiated with UV light (320-380 nm). In the presence of Ps, singlet oxygen was produced in the respiratory medium but neither the respiration nor the oxidative phosphorylation were affected. On the contrary the hydrophobic derivative TMP impaired the respiration with rapid uncoupling of oxidative phosphorylation as did Hp. The ineffectiveness of Ps as well as the effectiveness of TMP vs. Hp is explained on the basis of photophysical properties of the molecules and their partition coefficient. These results may indicate that, in the photochemiotherapy of skin diseases, furocoumarins can drive photodynamic reactions at various subcellular levels according to their hydrophobicity.     

EFFECTS OF RHODAMINE 123 IN THE DARK AND AFTER IRRADIATION ON MITOCHONDRIAL ENERGY METABOLISM

Isolated rat liver mitochondria have been used to study the mechanism of toxicity of Rhodamine 123 (Rho 123) in the dark and after irradiation with visible light. We report an inhibition of adenosine 5'-diphosphate phosphorylation which is increased after illumination. In the dark, the first steps of the phosphorylation process (i.e. the entry of substrates into the matrix, the electron transport to oxygen and the creation of the proton gradient) as well as ATPase activity are not significantly perturbed at Rho 123 concentration below 10 micrograms/mL. In contrast, the movements of the phosphate compounds are drastically impaired. Irradiation strengthens the detrimental effects in an oxygen dependent process. The nature of the noxious transient species is not clearly established, but it is suggested that singlet oxygen could be responsible for the observed damage.     

Photosensitization of isolated mitochondria by hematoporphyrin derivative (Photofrin): effects on bioenergetics.

Isolated rat liver mitochondria were incubated in the presence of 6 micrograms/ml of Photofrin and irradiated at the wavelength of 365 nm. After 45 s irradiation (30 W/m2), coupling defined as stimulation of respiration by externally added adenosine 5'-diphosphate (ADP) is totally lost. In contrast, membrane potential created by addition of succinate or adenosine 5'-triphosphate (ATP) is only slightly affected. Similarly, the ADP/O ratio is not modified after 20 s irradiation. These data suggest that modification of the mitochondrial membrane potential is not a primary event after irradiation.     

PHOTOSENSITIZATION OF MITOCHONDRIA BY LIPOSOME-BOUND PORPHYRINS

We have compared the photodynamic activities of hematoporphyrin (HP) and protoporphyrin (PP) on isolated rat liver mitochondria by measuring the decline of the respiratory control ratio (RCR) after irradiation at 365 nm. Before addition to the respiratory mcdium, the dyes were dissolved in phosphate-buffered saline (PBS) or incorporated into unilamellar liposomes of dipalmitoyl-phosphatidylcholine (DPPC), sometimes enriched with cholesterol (Chol) or cardiolipin (Card), which are naturally present in mitochondrial membranes. Chol and especially Card strongly increase the porphyrin uptake by mitochondria. In all experimental conditions, PP is taken up by mitochondria to a higher extent than HP. Nevertheless, under conditions giving the same amount of mitochondriabound dye, HP is a morc efficient photosensitizer than PP. As the efficiency of singlet oxygen production has been shown to be equivalent for the two porphyrins in monomeric state, the resulting photobiological effects are explained in terms of different localization of HP and PP in the mitochondrial membrancs. In particular, HP preferentially localizes in the protein-rich polar domains of the inner mitochondrial membrane, whereas PP dissolvcs in the lipid regions of the mcmbrancs.