S,S-Chiral Linker Induced U Shape with a Syn-facial Sensitizer and Photocleavable Ethene Group.

There is a major need for light-activated materials for the release of sensitizers and drugs. Considering the success of chiral columns for the separation of enantiomer drugs, we synthesized an S,S-chiral linker system covalently attached to silica with a sensitizer ethene near the silica surface. First, the silica surface was modified to be aromatic rich, by replacing 70% of the surface groups with (3-phenoxypropyl)silane. We then synthesized a 3-component conjugate [chlorin sensitizer, S,S-chiral cyclohexane and ethene building blocks] in 5 steps with a 13% yield, and covalently bound the conjugate to the (3-phenoxypropyl)silane-coated silica surface. We hypothesized that the chiral linker would increase exposure of the ethene site for enhanced 1 O2 -based sensitizer release. However, the chiral linker caused the sensitizer conjugate to adopt a U shape due to favored 1,2-diaxial substituent orientation; resulting in a reduced efficiency of surface loading. Further accentuating the U shape was π-π stacking between the (3-phenoxypropyl)silane and sensitizer. Semiempirical calculations and singlet oxygen luminescence data provided deeper insight into the sensitizer's orientation and release. This study has lead to insight on modifications of surfaces for drug photorelease and can help lead to the development of miniaturized photodynamic devices.

Superhydrophobic Photosensitizers: Airborne 1O2 Killing of an in Vitro Oral Biofilm at the Plastron Interface.

Singlet oxygen is a potent agent for the selective killing of a wide range of harmful cells; however, current delivery methods pose significant obstacles to its widespread use as a treatment agent. Limitations include the need for photosensitizer proximity to tissue because of the short (3.5 µs) lifetime of singlet oxygen in contact with water; the strong optical absorption of the photosensitizer, which limits the penetration depth; and hypoxic environments that restrict the concentration of available oxygen. In this article, we describe a novel superhydrophobic singlet oxygen delivery device for the selective inactivation of bacterial biofilms. The device addresses the current limitations by: immobilizing photosensitizer molecules onto inert silica particles; embedding the photosensitizer-containing particles into the plastron (i.e. the fluid-free space within a superhydrophobic surface between the solid substrate and fluid layer); distributing the particles along an optically transparent substrate such that they can be uniformly illuminated; enabling the penetration of oxygen via the contiguous vapor space defined by the plastron; and stabilizing the superhydrophobic state while avoiding the direct contact of the sensitizer to biomaterials. In this way, singlet oxygen generated on the sensitizer-containing particles can diffuse across the plastron and kill bacteria even deep within the hypoxic periodontal pockets. For the first time, we demonstrate complete biofilm inactivation (>5 log killing) of Porphyromonas gingivalis, a bacterium implicated in periodontal disease using the superhydrophobic singlet oxygen delivery device. The biofilms were cultured on hydroxyapatite disks and exposed to active and control surfaces to assess the killing efficiency as monitored by colony counting and confocal microscopy. Two sensitizer particle types, a silicon phthalocyanine sol-gel and a chlorin e6 derivative covalently bound to fluorinated silica, were evaluated; the biofilm killing efficiency was found to correlate with the amount of singlet oxygen detected in separate trapping studies. Finally, we discuss the applications of such devices in the treatment of periodontitis.

Experimental and DFT Computational Insight into Nitrosamine Photochemistry-Oxygen Matters.

A nitrosamine photooxidation reaction is shown to generate a peroxy intermediate by experimental physical-organic methods. The irradiation of phenyl and methyl-substituted nitrosamines in the presence of isotopically labeled 18-oxygen revealed that an O atom was trapped from a peroxy intermediate to trimethylphosphite or triphenylphosphine, or by nitrosamine itself, forming two moles of nitramine. The unstable peroxy intermediate can be trapped at low temperature in postphotolyzed solution in the dark. Chemiluminescence was also observed upon thermal decomposition of the peroxy intermediate, that is, when a postphotolysis low-temperature solution is brought up to room temperature. A DFT study provides tentative information for cyclic nitrogen peroxide species on the reaction surface.

Synthesis, Photophysics and PDT Evaluation of Mono-, Di-, Tri- and Hexa-PEG Chlorins for Pointsource Photodynamic Therapy.

Pointsource photodynamic therapy (PSPDT) is a newly developed fiber optic method aimed at the delivery of photosensitizer, light and oxygen to a diseased site. Because of a need for developing photosensitizers with desirable properties for PSPDT, we have carried out a synthetic, photophysical and phototoxicity study on a series of PEGylated sensitizers. Chlorin and pheophorbide sensitizers were readily amenable to our synthetic PEGylation strategy to reach triPEG and hexaPEG galloyl pheophorbides and mono-, di-, triPEG chlorins. On screening these PEG sensitizers, we found that increasing the number of PEG groups, except for hexaPEGylation, increases phototoxicity. We found that three PEG groups but not less or more were optimal. Of the series tested, a triPEG gallyol pheophorbide and a triPEG chlorin were the most efficient at generating singlet oxygen, and produced the highest phototoxicity and lowest dark toxicity to Jurkat cells. A detailed kinetic analysis of the PEGylated sensitizers in solution and cell culture and media is also presented. The data provide us with steps in the development of PSPDT to add to the PDT tools we have in general.

On the in vivo photochemical rate parameters for PDT reactive oxygen species modeling.

Photosensitizer photochemical parameters are crucial data in accurate dosimetry for photodynamic therapy (PDT) based on photochemical modeling. Progress has been made in the last few decades in determining the photochemical properties of commonly used photosensitizers (PS), but mostly in solution or in vitro. Recent developments allow for the estimation of some of these photochemical parameters in vivo. This review will cover the currently available in vivo photochemical properties of photosensitizers as well as the techniques for measuring those parameters. Furthermore, photochemical parameters that are independent of environmental factors or are universal for different photosensitizers will be examined. Most photosensitizers discussed in this review are of the type II (singlet oxygen) photooxidation category, although type I photosensitizers that involve other reactive oxygen species (ROS) will be discussed as well. The compilation of these parameters will be essential for ROS modeling of PDT.

Type I and Type II Photosensitized Oxidation Reactions: Guidelines and Mechanistic Pathways.

Here, 10 guidelines are presented for a standardized definition of type I and type II photosensitized oxidation reactions. Because of varied notions of reactions mediated by photosensitizers, a checklist of recommendations is provided for their definitions. Type I and type II photoreactions are oxygen-dependent and involve unstable species such as the initial formation of radical cation or neutral radicals from the substrates and/or singlet oxygen (1 O21 ∆g ) by energy transfer to molecular oxygen. In addition, superoxide anion radical (O2·-) can be generated by a charge-transfer reaction involving O2 or more likely indirectly as the result of O2 -mediated oxidation of the radical anion of type I photosensitizers. In subsequent reactions, O2·- may add and/or reduce a few highly oxidizing radicals that arise from the deprotonation of the radical cations of key biological targets. O2·- can also undergo dismutation into H2 O2 , the precursor of the highly reactive hydroxyl radical (·OH) that may induce delayed oxidation reactions in cells. In the second part, several examples of type I and type II photosensitized oxidation reactions are provided to illustrate the complexity and the diversity of the degradation pathways of mostly relevant biomolecules upon one-electron oxidation and singlet oxygen reactions.

"Ene" Reactions of Singlet Oxygen at the Air-Water Interface.

Prenylsurfactants [(CH3)2C═CH(CH2)nSO3(-) Na(+) (n = 4, 6, or 8)] were designed to probe the "ene" reaction mechanism of singlet oxygen at the air-water interface. Increasing the number of carbon atoms in the hydrophobic chain caused an increase in the regioselectivity for a secondary rather than tertiary surfactant hydroperoxide, arguing for an orthogonal alkene on water. The use of water, deuterium oxide, and H2O/D2O mixtures helped to distinguish mechanistic alternatives to homogeneous solution conditions that include dewetting of the π bond and an unsymmetrical perepoxide transition state in the hydroperoxide-forming step. The prenylsurfactants and a photoreactor technique allowed a certain degree of interfacial control of the hydroperoxidation reaction on a liquid support, where the oxidant (airborne (1)O2) is delivered as a gas.

Synthesis of a poly(ethylene glycol) galloyl sensitizer tip for an 'all-in-one' photodynamic device.

This paper describes the synthesis of a specialized silica tip for an optical fiber device capable of delivering all components necessary for photodynamic therapy. Oxygen, light and a cleavable tripolyethylene glycol (PEG)-galloyl pheophorbide sensitizer are simultaneously delivered by the silica tip, where the tip was synthesized in six steps. A comparison of synthetic steps to reach PEGylated sensitizers bound to fluorinated silica and a previously reported Teflon/polyvinyl alcohol (PVA) nanocomposite ( Ghosh, G. et al. J. Phys. Chem. B 2015, 119, 4155- 4164) was made. The hydrolytic stability of the attached PEGs and the extent to which the PEG groups enhance solubility will also be discussed. The new triPEG-galloyl sensitizer has the potential for use in intraoperative pointsource photodynamic therapy which aims for precision treatment of residual disease. Schematic of the synthesis of a photoactive silica surface. It is composed of fluorinated silica connected to a photo-releasable sensitizer with short-chain PEGs.

Using Singlet Oxygen to Synthesize Natural Products and Drugs.

This Review describes singlet oxygen ((1)O2) in the organic synthesis of targets on possible (1)O2 biosynthetic routes. The visible-light sensitized production of (1)O2 is not only useful for synthesis; it is extremely common in nature. This Review is intended to draw a logical link between flow and batch reactions-a combination that leads to the current state of (1)O2 in synthesis.

Synthesis and antiproliferative properties of a new ceramide analog of varacin.

A benzopentasulfane was synthesized in 8 steps with a ceramide attached through an amide bond to the 7-position of the heterocycle structure. The anticancer activity of this synthetic ceramide-benzopolysulfane drug conjugate was analyzed against five human cancer cell lines MDA-MB-231 (breast), DU145 (prostate), MIA PaCa-2 (pancreas), HeLa (cervix), and U251 (glioblastoma). The ceramide-benzopolysulfane conjugate had IC50 values ranging from 10 to >20 µM with complete cell killing at 12.5 µM for MDA-MB-231 and 20 µM for DU145 and HeLa cells. The ceramide-benzopolysulfane conjugate had IC50 values 1.8 and 4.0 times lower than a PEG benzopolysulfane, N-(2-(2-(2-methoxyethoxy)ethoxy)ethyl)benzo[f][1,2,3,4,5]-pentathiepine-7-carboxamide, for MDA-MB-231 and DU145 cells, respectively. The parent "unsubstituted" benzopolysulfane, o-C6H4S5, had IC50 values 4.2 times lower and 2.7 times higher than the ceramide benzopolysulfane for MDA-MB-231 and DU145 cells, respectively. The results indicate that the polysulfur linkage is needed for activity since benzenedithiol, o-C6H4(SH)2, had IC50 values greater than 30 µM with little effect on MDA-MB-231 and DU145 cells. Thus, to account for the bioactivity, a bimolecular reaction of cellular thiol with the ceramide benzopolysulfane is a proposed followed by thiozone (S3) extrusion.

Fluorinated Photodynamic Therapy Device Tips and their Resistance to Fouling for In Vivo Sensitizer Release.

We describe progress on a one-step photodynamic therapy (PDT) technique that is simple: device tip delivery of sensitizer, oxygen and light simultaneously. Control is essential for their delivery to target sites to generate singlet oxygen. One potential problem is the silica device tip may suffer from biomaterial fouling and the pace of sensitizer photorelease is slowed. Here, we have used biomaterial (e.g. proteins, cells, etc.) from SQ20B head and neck tumors and whole blood for an assessment of fouling of the silica tips by adsorption. It was shown that by exchanging the native silica tip for a fluorinated tip, a better nonstick property led to an increased sensitizer output by ~10%. The fluorinated tip gave a sigmoidal photorelease where singlet oxygen is stabilized to physical quenching, whereas the native silica tip with unprotected SiO-H groups gave a slower (pseudolinear) photorelease. A further benefit from fluorinated silica is that 15% less biomaterial adheres to its surface compared to native silica based on a bicinchoninic acid assay (BCA) and X-ray photoelectron spectroscopy (XPS) measurements. We discuss how the fluorination of the device tip increases biofouling resistance and can contribute to a new pointsource PDT tool.

Mechanism of Photochemical O-Atom Exchange in Nitrosamines with Molecular Oxygen.

The detection of an oxygen-atom photoexchange process of N-nitrosamines is reported. The photolysis of four nitrosamines (N-nitrosodiphenylamine 1, N-nitroso-N-methylaniline 2, N-butyl-N-(4-hydroxybutyl)nitrosamine 3, and N-nitrosodiethylamine 4) with ultraviolet light was examined in an (18)O2-enriched atmosphere in solution. HPLC/MS and HPLC-MS/MS data show that (18)O-labeled nitrosamines were generated for 1 and 2. In contrast, nitrosamines 3 and 4 do not exchange the (18)O label and instead decomposed to amines and/or imines under the conditions. For 1 and 2, the (18)O atom was found not to be introduced by moisture or by singlet oxygen [(18)((1)O2 (1)Δg)] produced thermally by (18)O-(18)O labeled endoperoxide of N,N'-di(2,3-hydroxypropyl)-1,4-naphthalene dipropanamide (DHPN(18)O2) or by visible-light sensitization. A density functional theory study of the structures and energetics of peroxy intermediates arising from reaction of nitrosamines with O2 is also presented. A reversible head-to-tail dimerization of the O-nitrooxide to the 1,2,3,5,6,7-hexaoxadiazocane (30 kcal/mol barrier) with extrusion of O═(18)O accounts for exchange of the oxygen atom label. The unimolecular cyclization of O-nitrooxide to 1,2,3,4-trioxazetidine (46 kcal/mol barrier) followed by a retro [2 + 2] reaction is an alternative, but higher energy process. Both pathways would require the photoexcitation of the nitrooxide.

Photoactive fluoropolymer surfaces that release sensitizer drug molecules.

We describe a physical-organic study of two fluoropolymers bearing a photoreleasable PEGylated photosensitizer that generates (1)O2((1)Δg) [chlorin e6 methoxy tri(ethylene glycol) triester]. The surfaces are Teflon/poly(vinyl alcohol) (PVA) nanocomposite and fluorinated silica. The relative efficiency of these surfaces to photorelease the PEGylated sensitizer [shown previously to be phototoxic to ovarian cancer cells (Kimani, S. et al. J. Org. Chem 2012, 77, 10638)] was slightly higher for the nanocomposite. In the presence of red light and O2, (1)O2 is formed, which cleaves an ethene linkage to liberate the sensitizer in 68-92% yield. The fluoropolymers were designed to deal with multiple problems. Namely, their success relied not only on high O2 solubility and drug repellency but also on the C-F bonds, which physically quench little (1)O2, for singlet oxygen's productive use away from the surface. The results obtained here indicate that Teflon-like surfaces have potential uses in delivering sensitizer and singlet oxygen for applications in tissue repair and photodynamic therapy (PDT).

Air-Water Interface Effects on the Regioselectivity of Singlet Oxygenations of a Trisubstituted Alkene.

The regioselective synthesis of allylic hydroperoxide sulfonates by singlet oxygenation at the air-water interface has been found to depend on the concentration of the alkene sulfonate and added calcium salt. The regioselectivity is proposed to originate from an orthogonal alkene relative to the water surface for preferential methyl hydrogen abstraction by airborne singlet oxygen in an ene reaction. The findings hint that the air-water interface is a locale for synthetic reactions.

"Pointsource" delivery of a photosensitizer drug and singlet oxygen: eradication of glioma cells in vitro.

We describe a pointsource sensitizer-tipped microoptic device for the eradication of glioma U87 cells. The device has a mesoporous fluorinated silica tip which emits singlet oxygen molecules and small quantities of pheophorbide sensitizer for additional production of singlet oxygen in the immediate vicinity. The results show that the device surges in sensitizer release and photokilling with higher rates about midway through the reaction. This was attributed to a self-amplified autocatalytic reaction where released sensitizer in the extracellular matrix provides positive feedback to assist in the release of additional sensitizer. The photokilling of the glioma cells was analyzed by global toxicity and live/dead assays, where a killing radius around the tip with ~0.3 mm precision was achieved. The implication of these results for a new PDT tool of hard-to-resect tumors, e.g. in the brain, is discussed.

Theoretical study of the reaction formalhydrazone with singlet oxygen. Fragmentation of the C=N bond, ene reaction and other processes.

Photobiologic and synthetic versatility of hydrazones has not yet been established with (1)O2 as a route to commonly encountered nitrosamines. Thus, to determine whether the "parent" reaction of formalhydrazone and (1)O2 leads to facile C=N bond cleavage and resulting nitrosamine formation, we have carried out CCSD(T)//DFT calculations and analyzed the energetics of the oxidation pathways. A [2 + 2] pathway occurs via diradicals and formation of 3-amino-1,2,3-dioxazetidine in a 16 kcal/mol(-1) process. Reversible addition or physical quenching of (1)O2 occurs either on the formalhydrazone carbon for triplet diradicals at 2-3 kcal mol(-1), or on the nitrogen (N(3)) atom forming zwitterions at ~15 kcal/mol(-1), although the quenching channel by charge-transfer interaction was not computed. The computations also predict a facile conversion of formalhydrazone and (1)O2 to hydroperoxymethyl diazene in a low-barrier 'ene' process, but no 2-amino-oxaziridine-O-oxide (perepoxide-like) intermediate was found. A Benson-like analysis (group increment calculations) on the closed-shell species are in accord with the quantum chemical results.