Santi Nonell

Aromatic ketones as standards for singlet molecular oxygen O2(1Δg) photosensitization. Time-resolved photoacoustic and near-IR emission studies

Abstract Time-resolved near-IR emission and optoacoustic calorimetry studies were carried out on the aromatic ketones phenalenone, benzanthrone, 4-phenylbenzophenone and the benzophenone-naphthalene (0.1 M) system in order to assess their adoption as solvent-independent standards for singlet molecular oxygen O 2 ( 1 Δ g ) photosensitization. All compounds show quantum yields of O 2 ( 1 Δ g ) production (FΔ) in the range 0.9–1 in cyclohexane. Increasing solvent polarity or protic character reduces the FΔ values for all sensitizers except phenalenone. Laser-induced optoacoustic calorimetry was used to obtain the absolute FΔ values for the latter compound by applying both maximum amplitude and deconvolution methods. The former yields highly precise results (3%–5% uncertainty) and has been chosen for standardization purposes. The deconvolution method yields both kinetic and quantum yield data, albeit with lower precision (10%–15% uncertainty).

Porphycenes: Facts and Prospects in Photodynamic Therapy of Cancer

The photodynamic process induces cell damage and death by the combined effect of a photosensitizer (PS), visible light, and molecular oxygen, which generate singlet oxygen ((1)O(2)) and other reactive oxygen species that are responsible for cytotoxicity. The most important application of this process with increasing biomedical interest is the photodynamic therapy (PDT) of cancer. In addition to hematoporphyrin-based drugs, 2nd generation PSs with better photochemical properties are now studied using cell cultures, experimental tumors and clinical trials. Porphycene is a structural isomer of porphyrin and constitutes an interesting new class of PS. Porphycene derivatives show higher absorption than porphyrins in the red spectral region (lambda > 600 nm, epsilon > 50000 M-(1)cm(-1)) owing to the lower molecular symmetry. Photophysical and photobiological properties of porphycenes make them excellent candidates as PSs, showing fast uptake and diverse subcellular localizations (mainly membranous organelles). Several tetraalkylporphycenes and the tetraphenyl derivative (TPPo) induce photodamage and cell death in vitro. Photodynamic treatments of cultured tumor cells with TPPo and its palladium(II) complex induce cytoskeletal changes, mitotic blockage, and dose-dependent apoptotic or necrotic cell death. Some pharmacokinetic and phototherapeutic studies on experimental tumors after intravenous or topical application of lipophilic alkyl-substituted porphycene derivatives are known. Taking into account all these features, porphycene PSs should be very useful for PDT of cancer and other biomedical applications.

[4] Time-resolved singlet oxygen detection

Publisher Summary Techniques for the detection of singlet oxygen [O 2 ( 1 Δ g )] are increasingly implemented in many laboratories, reflecting the growing awareness that O 2 ( 1 Δ g ) plays crucial roles in several photoexcited systems. Time-resolved detection techniques have been used for (1) the identification of O 2 ( 1 Δ g ), (2) the measurement of quantum yields of O 2 ( 1 Δ g ) production in photosensitized processes, Φ Δ , and (3) the determination of rate constant for the interaction of O 2 ( 1 Δ g ) with substrates. This chapter describes the methods used most commonly to attain the just-described goals associated with the technique of time-resolved near-infrared detection of O 2 ( 1 Δ g ) phosphorescence at 1.27μm (TRNIR). Focus is placed on laboratory procedures rather than on techniques. Only methods for homogeneous solutions are discussed. Clues are given for extending them to more complex systems, but the number of possible situations precludes general statements. It is restricted to systems in which O 2 ( 1 Δ g ) is produced by photosensitization, thus excluding chemical or enzymatic production.

Photodynamic inactivation of Acinetobacter baumannii using phenothiazinium dyes: in vitro and in vivo studies.

Phenothiazinium dyes have been reported to be effective photosensitizers inactivating a wide range of microorganisms in vitro after illumination with red light. However, their application in vivo has not extensively been explored. This study evaluates the bactericidal activity of phenothiazinium dyes against multidrug‐resistant Acinetobacter baumannii both in vitro and in vivo.

Cationic Porphycenes as Potential Photosensitizers for Antimicrobial Photodynamic Therapy

Structures of typical photosensitizers used in antimicrobial photodynamic therapy are based on porphyrins, phthalocyanines, and phenothiazinium salts, with cationic charges at physiological pH values. However, derivatives of the porphycene macrocycle (a structural isomer of porphyrin) have barely been investigated as antimicrobial agents. Therefore, we report the synthesis of the first tricationic water-soluble porphycene and its basic photochemical properties. We successfully tested it for in vitro photoinactivation of different Gram-positive and Gram-negative bacteria, as well as a fungal species (Candida) in a drug-dose and light-dose dependent manner. We also used the cationic porphycene in vivo to treat an infection model comprising mouse third degree burns infected with a bioluminescent methicillin-resistant Staphylococcus aureus strain. There was a 2.6-log(10) reduction (p < 0.001) of the bacterial bioluminescence for the PDT-treated group after irradiation with 180 J·cm(-2) of red light.

Singlet oxygen in Escherichia coli: New insights for antimicrobial photodynamic therapy

Antimicrobial photodynamic therapy is an emerging treatment for bacterial infections that is becoming increasingly more attractive because of its effectiveness and unlikelihood of inducing bacterial resistance. However, there is limited knowledge about the localization of the photoactive drug in the bacteria and about the details of production of the main cytotoxic species, singlet oxygen. This article describes a combination of spectroscopic and time-resolved photophysical techniques that provide such information for a cationic porphyrin photosensitizer in gram-negative Escherichia coli bacteria. Our results reveal a double localization of the photosensitizer, inside (bound to the nucleic acids) and outside (bound to the cell wall) of the E. coli cells. Singlet oxygen is produced at both sites and is able to cross the cell wall.

Singlet Oxygen Generation by the Genetically Encoded Tag miniSOG

The genetically encodable fluorescent tag miniSOG is expected to revolutionize correlative light- and electron microscopy due to its ability to produce singlet oxygen upon light irradiation. The quantum yield of this process was reported as ΦΔ = 0.47 ± 0.05, as derived from miniSOGs ability to photooxidize the fluorescent probe anthracene dipropionic acid (ADPA). In this report, a significantly smaller value of ΦΔ = 0.03 ± 0.01 is obtained by two methods: direct measurement of its phosphorescence at 1275 nm and chemical trapping using uric acid as an alternative probe. We present insight into the photochemistry of miniSOG and ascertain the reasons for the discrepancy in ΦΔ values. We find that miniSOG oxidizes ADPA by both singlet oxygen-dependent and -independent processes. We also find that cumulative irradiation of miniSOG increases its ΦΔ value ~10-fold due to a photoinduced transformation of the protein. This may be the reason why miniSOG outperforms other fluorescent proteins reported to date as singlet oxygen generators.

Kinetics of singlet oxygen photosensitization in human skin fibroblasts

The roles played by singlet oxygen ((1)O(2)) in photodynamic therapy are not fully understood yet. In particular, the mobility of (1)O(2) within cells has been a subject of debate for the last two decades. In this work, we report on the kinetics of (1)O(2) formation, diffusion, and decay in human skin fibroblasts. (1)O(2) has been photosensitized by two water-soluble porphyrins targeting different subcellular organelles, namely the nucleus and lysosomes, respectively. By recording the time-resolved near-IR phosphorescence of (1)O(2) and that of its precursor the photosensitizers triplet state, we find that the kinetics of singlet oxygen formation and decay are strongly dependent on the site of generation. (1)O(2) photosensitized in the nucleus is able to escape out of the cells while (1)O(2) photosensitized in the lysosomes is not. Despite showing a lifetime in the microsecond time domain, (1)O(2) decay is largely governed by interactions with the biomolecules within the organelle where it is produced. This observation may reconcile earlier views that singlet oxygen-induced photodamage is highly localized, while its lifetime is long enough to diffuse over long distances within the cells.

Photoantimicrobials—are we afraid of the light?

Summary Although conventional antimicrobial drugs have been viewed as miraculous cure-alls for the past 80 years, increasing antimicrobial drug resistance requires a major and rapid intervention. However, the development of novel but still conventional systemic antimicrobial agents, having only a single mode or site of action, will not alleviate the situation because it is probably only a matter of time until any such agents will also become ineffective. To continue to produce new agents based on this notion is unacceptable, and there is an increasing need for alternative approaches to the problem. By contrast, light-activated molecules called photoantimicrobials act locally via the in-situ production of highly reactive oxygen species, which simultaneously attack various biomolecular sites in the pathogenic target and therefore offer both multiple and variable sites of action. This non-specificity at the target circumvents conventional mechanisms of resistance and inhibits the development of resistance to the agents themselves. Photoantimicrobial therapy is safe and easy to implement and, unlike conventional agents, the activity spectrum of photoantimicrobials covers bacteria, fungi, viruses, and protozoa. However, clinical trials of these new, truly broad-spectrum, and minimally toxic agents have been few, and the funding for research and development is almost non-existent. Photoantimicrobials constitute one of the few ways forward through the morass of drug-resistant infectious disease and should be fully explored. In this Personal View, we raise awareness of the novel photoantimicrobial technologies that offer a viable alternative to conventional drugs in many relevant application fields, and could thus slow the pace of resistance development.

Solvent influence on the kinetics of the photodynamic degradation of trolox, a water-soluble model compound for vitamin E

The kinetics of the aerobic dye-sensitized photo-oxidation of 6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid (Trolox), a water-soluble model compound for vitamin E, has been studied in aqueous solutions of pH values 6 and 12 and in organic media of different polarity. Experimental evidence indicates that the process largely occurs through a Trolox-singlet molecular oxygen O2(1Δg) interaction, the photo-oxidation rates increasing with the pH value and the polarity of the medium. Low polarity solvents favour the physical quenching of O2(1Δg) by Trolox, as already observed in other tocopherols. High polarity solvents drive the interaction predominantly towards a reactive pathway that yields the same substituted p-benzoquinone that can be obtained by thermal oxidation of Trolox. Overall rate constants for the quenching of O2(1Δg) by Trolox (determined by time-resolved phosphorescence spectroscopy) and reactive rate constants (obtained by relative methods) are both in the range 107−108 M−1s−1. On the basis of these and their results and their comparison with previous results on the photo-oxidation of tocopherol derivatives, the role of the solvent in the behaviour of vitamin E is discussed.