Jerry C. Bommer

Photophysical Properties and Antibacterial Activity of Meso-substituted Cationic Porphyrins¶

A series of derivatives of 5,10,15,20‐tetrakis‐(4‐N‐methylpyridyl)‐porphine, where one N‐methyl group was replaced by a hydrocarbon chain ranging from C6 to C22, were characterized for their photophysical and photosensitizing properties. The absorption and fluorescence features of the various compounds in neutral aqueous solutions were typical of largely monomeric porphyrins, with the exception of the C22 derivative, which appeared to be extensively aggregated. This was confirmed by the very low triplet quantum yield and lifetime of the C22 derivative as compared with 0.2–0.7 quantum yields and 88–167 μs lifetimes for the other porphyrins. The photophysical properties and photosensitizing activity toward N‐acetyl‐l‐tryptophanamide of the C22 porphyrin became comparable to those typical of the other derivatives in 2% aqueous sodium dodecyl sulfate, where the C22 compound is fully monomerized. All the porphyrin derivatives exhibited at micromolar concentrations photoinactivation activity against both Staphylococcus aureus and Escherichia coli, even though the gram‐negative bacteria were markedly less photosensitive. The photosensitizing efficiency was influenced by (1) the amount of cell‐bound porphyrin, which increased with increasing length of the hydrocarbon chain; and (2) the tendency to undergo partial aggregation in the cell, which seems to be especially important for the C22 derivative.

Photosensitizing properties of mono-l-aspartyl chlorin e6 (NPe6): A candidate sensitizer for the photodynamic therapy of tumors

There is a large amount of interest in chlorins as photosensitizers for the photodynamic therapy of tumors because of their strong absorption in the red, where light penetration into mammalian tissues is efficient. Mono-L-aspartyl chlorin e6 (NPe6), in phosphate buffer of pH 7.4, had absorption peaks at 400 and 654 nm with molar absorption coefficients of 180,000 and 40,000 M-1 cm-1 respectively. In buffer, the NPe6 triplet had a peak at 440 nm and a lifetime under argon of approximately 300 microseconds. The triplet was efficiently quenched by ground state oxygen (kQ = 1.9 x 10(9) M-1 s-1) with the formation of singlet oxygen, as identified by its near infrared luminescence. The quantum yield of singlet oxygen production was 0.77. A number of substrates were efficiently photo-oxidized by NPe6, including furfuryl alcohol, cysteine, histidine, tryptophan and human serum albumin. These reactions were efficiently inhibited by azide (which did not quench NPe6 triplets), indicating that they are probably mediated by singlet oxygen. Thus, NPe6 has a desirable array of photoproperties for a sensitizer to be used in the clinical photodynamic therapy of tumors.

Zinc Tetrasulphophthalocyanine as a Photodynamic Sensitizer for Biomolecules

Monomeric zinc tetrasulphophthalocyanine in aqueous buffer is an effective sensitizer for the photo-oxidation of amino acids and nucleic acid bases and for the photodynamic inactivation of the enzyme, lysozyme; these reactions appear to be mediated by singlet oxygen.

A comparison of the photoproperties of zinc phthalocyanine and zinc naphthalocyanine tetrasulfonates : model sensitizers for the photodynamic therapy of tumors

Abstract Phthalo- and naphthaocyanines are of interest as sensitizers for the photodynamic therapy of tumors because of their strong absorption in the 680 and 760 nm ranges respectively. Both zinc phthalocyanine and naphthalocyanine tetrasulfonates (ZnPcS 4 and ZnNcS 4 ) were aggregated and photochemically inactive in aqueous buffer of pH 7.4, while in 10 mM cetyl pyridinium chloride in buffer they were monomeric and active. Therefore all these studies were carried out using the buffered detergent. The triplet lifetimes of ZnPcS 4 and ZnNcS 4 under argon were 490 and 110 μs respectively, with oxygen bimolecular quenching constants of 4.2 × 10 9 and 2.0 × 10 8 M −1 s −1 respectively. Triplet decay curves in argon, air and 100% oxygen were first order, suggesting that there was little back reaction of the triplet states with oxygen as has been observed with some naphthalocyanines. The quantum yield of singlet oxygen generation by ZnPcS 4 was 0.70 and that for ZnNcS 4 was 0.25. Both compounds sensitized the photo-oxidation of furfuryl alcohol, cysteine, histidine, methionine, tryptophan, tyrosine and guanosine; ZnPcS 4 was three times more efficient than ZnNcS 4 . These reactions were 50% inhibited by about 0.5 mM azide, suggesting the involvement of singlet oxygen. Both sensitizers photobleached on illumination, with quantum yields of 1.7 × 10 −5 for ZnPcS 4 and 4.2 × 10 −3 for ZnNcS 4 .

PHOTOBLEACHING OF MONO‐l‐ASPARTYL CHLORIN e6 (NPe6): A CANDIDATE SENSITIZER FOR THE PHOTODYNAMIC THERAPY OF TUMORS

Abstract— Most sensitizers used for the photodynamic therapy (PDT) of tumors photobleach on illumination. Thus, it is of interest to examine the photobleaching behavior of new sensitizers proposed for use in PDT. This report surveys the quantum yields and kinetics of the photobleaching of mono‐l‐aspartyl chlorin e6 (NPe6), a hydrophilic chlorin that has many of the photoproperties desirable in a sensitizer for clinical PDT. It is a very effective sensitizer for the PDT of several types of model tumors in animals and is now in Phase I clinical trials. The quantum yield of NPe6 photobleaching in pH 7.4 phosphate buffer in air was 8.2 × 10−4; this is greater than the yields for typical porphyrin photosensitizers. For example, the yields for hematoporphyrin and uroporphyrin are 4.7 × 10 5 and 2.8 × 10−5, respectively. The yield decreased significantly in organic solvents of low dielectric constant. The Sn derivative of NPe6 was more light stable than NPe6 (yield = 5.7 × 10 −6), while the Zn derivative was more sensitive (yield = 1.9 × 10−2). Oxygen appeared to be necessary for the photobleaching of NPe6; however, bleaching was not inhibited by 100 mM azide, an efficient quencher of singlet oxygen. The photooxidizable substrates cysteine, dithiothreitol and furfuryl alcohol increased the quantum yield of photoblcaching two‐ to four‐fold, while the electron acceptor, met‐ronidazole, increased it almost six‐fold. Photobleaching yields for several other chlorins were also measured.

Cellular inhibition of microtubule assembly by photoactivated sulphonated meso-tetraphenylporphines

This work relates to studies on modes of phototoxicity by sulphonated mesotetraphenylporphines on cultured cells. Toxicity appears to be related to inhibition of microtubule function. Treatment of human cervix carcinoma cells of the line NHIK 3025 incubated for 18 h with meso-tetraphenylporphine sulphonates (TPPSn where n = 2a, 2o or 4) and exposed to light, inhibits multiplication for the first hours after light exposure, a significant fraction of the cells accumulating in mitosis. The maximal number of cells in mitosis after treatment (approximately 20%) is dependent on the fluence but is similar for all three photosensitizers. For the first hours after treatment the mitotic cells were always mainly in metaphase; mainly seen as c-metaphases and three-group metaphases. During this time anaphase and telophase cells were absent or greatly reduced in number. Indirect immunofluorescence staining of beta-tubulin showed that the spindle apparatus of mitotic cells was perturbed in all cases. Results are presented which indicate that photoactivation of TPPSn located on the plasma membrane destroys microtubules in interphase cells and leads to arrest of the cells in mitosis. The localization of the dye which sensitizes the photoinduced perturbation of microtubules is further discussed.

ACTION SPECTRA OF PHTHALOCYANINES WITH RESPECT TO PHOTOSENSITIZATION OF CELLS

Human carcinoma cells (NHIK 3025 cells) and Chinese hamster cells (V79 cells) were incubated with AIPcS1, AIPcS2 and AIPcS4, phthalocyanines with different lipophilicity but with similar photochemical properties when in monomeric solutions. The absorption‐ and fluorescence spectra of the dyes in the cells were recorded as well as their action spectra with respect to sensitizing cells to photoinactivation. These spectra show that under the present conditions AIPcS1 is strongly aggregated in both cell lines; AIPcS2 is aggregated in V79 cells but much less so in NHIK 3025 cells. A main finding is that the shapes of the action spectra are similar to that of the fluorescence excitation spectra, but not to the absorption spectra, indicating that the photosensitizing effects of the dyes are mainly due to their monomeric fraction in the cells. AIPcS2 and AIPcS4 localize intracellularly mainly in lysosomes while AIPcS1 was found to be more diffusely distributed in cells. As measured per quantum of fluorescence emitted, AIPcS2 and AIPcS2 are more efficient sensitizers than AIPcS4. The difference in efficiency between AIPcS2 and AIPcS4 is supposedly due to a different localization pattern on the suborganelle level.

In Vitro Evaluation of the Antimicrobial Agent AquaFrin as a Bactericide and Selective Algicide for Use in Channel Catfish Aquaculture

Abstract Producers of pond-raised channel catfish Ictalurus punctatus in the southeastern United States can experience huge economic losses due to enteric septicemia of catfish (ESC) and columnaris disease and to the presence of certain odor-producing cyanobacteria in production ponds that result in “off-flavor” channel catfish. AquaFrin (lauryl methyl pyrifrin), a product currently being evaluated for commercial use, belongs to a class of porphyrins that have been shown to be very effective as antibacterial agents. As a preliminary step to assess the potential use of AquaFrin as a therapeutant against ESC and columnaris, we used a rapid bioassay to evaluate AquaFrin for antibacterial activity against the Gram-negative bacterium Edwardsiella ictaluri (etiological agent of ESC), two genotypes of the Gram-negative bacterium Flavobacterium columnare (etiological agent of columnaris disease), and the representative Gram-positive bacterium Staphylococcus aureus (American Type Culture Collection 29213) for comp...