Photosensitized biological processes, as applied in photodynamic therapy, are based on light-triggered generation of molecular singlet oxygen by a membrane-residing sensitizer. Most of the sensitizers currently used are hydrophobic or amphiphilic porphyrins and their analogs. The possible activity of the short-lived singlet oxygen is limited to the time it is diffusing in the membrane, before it emerges into the aqueous environment. In this paper we demonstrate the enhancement of the photosensitization process that is obtained by newly synthesized protoporphyrin derivatives, which insert their tetrapyrrole chromophore deeper into the lipid bilayer of liposomes. The insertion was measured by fluorescence quenching by iodide and the photosensitization efficiency was measured with 9,10-dimethylanthracene, a fluorescent chemical target for singlet oxygen. We also show that when the bilayer undergoes a melting phase transition, or when it is fluidized by benzyl alcohol, the sensitization efficiency decreases because of the enhanced diffusion of singlet oxygen. The addition of cholesterol or of dimyristoyl phosphatydilcholine to the bilayer moves the porphyrin deeper into the bilayer; however, the ensuing effect on the sensitization efficiency is different in these two cases. These results could possibly define an additional criterion for the choice and design of hydrophobic, membrane-bound photosensitizers.

Ammonium trichloro(dioxoethylene-O,O‘)tellurate (AS101) is an organotellurium(IV) compound that exhibits immunomodulation activity. In light of the unique Te(IV)−thiol chemistry, it was tested as a selective cysteine protease inhibitor. Although no inhibitory activity of serine-, metallo-, or aspartic proteases was observed, AS101 exhibited time- and concentration-dependent inactivation of cysteine proteases. The kinetic parameters of inactivation of papain were K i = 3.5 ± 2.0 μM and k i = (5.1 ± 0.4) × 10-2 min-1. The enzymatic activity could be recovered by treatment with thiols, indicating that the inactivation involves oxidation of the active-site thiol to a disulfide bond (Enz−S−S−R) or to a species containing a Te−S bond such as Enz−S−Te−S−R. Gel permeation chromatography established that the R group is a small molecule and excludes the possibility of dimerization of the enzyme itself. It was further established that some other Te(IV) derivatives could also inactivate cysteine proteases, while Te(VI) derivatives did not exhibit any such inhibitory activity. In order to understand the chemistry underlying the cysteine protease inactivation by AS101 and other organotellurium(IV) compounds, their interaction with the model compound cysteine was studied. While the Te(VI) derivatives did not interact with cysteine, all of the Te(IV) compounds interacted with 4 equiv of cysteine. The kinetics of this interaction is first order in Te and second order in thiol, yielding a third-order rate constant of ∼106 M-2 s-1, as determined for the interaction between AS101 with cysteine. The interactions between Te derivatives and cysteine in DMSO were followed by 125Te and 13C NMR. While Te(VI) compounds did not undergo any changes upon interaction with cysteine, on the basis of their 125Te NMR, the Te(IV) derivatives interacted with 4 equiv of cysteine, yielding new stable Te(IV) compounds. These compounds were tentatively designated as Te(cysteine)4 or its high-valence complex with other components in the reaction mixture. These results expand our understanding of tellurium chemistry and correlate well with its biological activity. Such knowledge can be applied for the development of novel biologically active tellurium compounds.

Photosensitization by porphyrins and other tetrapyrrole chromophores is used in biology and medicine to kill cells. This light-triggered generation of singlet oxygen is used to eradicate cancer cells in a process dubbed "photodynamic therapy," or PDT. Most photosensitizers are of amphiphilic character and they partition into cellular lipid membranes. The photodamage that they inflict to the host cell is mainly localized in membrane proteins. This photosensitized damage must occur in competition with the rapid diffusion of singlet oxygen through the lipid phase and its escape into the aqueous phase. In this article we show that the extent of damage can be modulated by employing modified hemato- and protoporphyrins, which have alkyl spacers of varying lengths between the tetrapyrrole ring and the carboxylate groups that are anchored at the lipid/water interface. The chromophore part of the molecule, and the point of generation of singlet oxygen, is thus located at a deeper position in the bilayer. The photosensitization efficiency was measured with 9,10-dimethylanthracene, a fluorescent chemical target for singlet oxygen. The vertical insertion of the sensitizers was assessed by two fluorescence-quenching techniques: by iodide ions that come from the aqueous phase; and by spin-probe-labeled phospholipids, that are incorporated into the bilayer, using the parallax method. These methods also show that temperature has a small effect on the depth when the membrane is in the liquid phase. However, when the bilayer undergoes a phase transition to the solid gel phase, the porphyrins are extruded toward the water interface as the temperature is lowered. These results, together with a previous publication in this journal, represent a unique and precedental case where the vertical location of a small molecule in a membrane has an effect on its membranal activity.

Zirconium complexes of two dianionic amine bis(phenolate) ligands have been synthesized, their X-ray structures solved, and their activity as hex-1-ene polymerization catalysts studied; upon treatment with B(C 6 F 5 ) 3 , an octahedral [ONNO]Zr(CH 2 Ph) 2 complex, having an extra N-donor group on a side arm, shows exceptionally high polymerization activity and yields a high molecular weight poly(hex-1-ene), whereas a related pentacoordinate [ONO]Zr(CH 2 Ph) 2 complex, having no extra donor group, shows only poor activity as a polymerization catalyst.

The internalization mechanism and subcellular distribution of hypericin (Hyp), hypericin tetrasulfonic acid (HypS4) and 1,3,4,6-tetrahydroxyhelianthrone (Hel) were studied in murine colon carcinoma CT26 cells, in protein-free medium or in the presence of serum proteins. The correlation between the extent of uptake of the sensitizers by cells that were incubated in the presence of different serum components, and the internalization mechanisms, was studied. The results indicate that sensitizer internalization may be a result of three mechanisms: partitioning, pinocytosis and endocytosis, and as a direct consequence is targeted to specific subcellular sites. While Hyp and Hel, the two lipophilic sensitizers, were localized in the endoplasmic reticulum after protein-free internalization, the hydrophilic HypS4 was localized in the cytoplasmic membrane and in lysosomes. An endolysosomal internalization route was revealed for Hyp and Hel under serum-enriched conditions showing lysosomal localization, as for HypS4. The lysosomal accumulation of Hyp-serum and specifically Hyp-LDL points to an endocytotic mechanism which is supported by its higher uptake parameter in an LDL-enriched medium, compared to the medium with 10% serum. The different uptake parameters of Hyp to cells, with or without serum, reflect the different mechanisms. Smaller differences in the uptake parameter for HypS4 reflect the distinction between partitioning and endocytosis, which, in this case, are both targeted to the lysosomes. The same uptake parameter of Hel to cells incubated in media with or without serum indicates the absence of the endocytotic mechanism. The interrelationship between subcellular targeting and photodynamic treatment was shown for the three sensitizers Hyp was found to be the most efficient sensitizer for PDT under our illumination protocol and it was dependent on internalization and localization sites.