Photosensitized Singlet Oxygen Research Articles

Synthesis and characterization of aryl-substituted BODIPY dyes displaying distinct solvatochromic singlet oxygen photosensitization efficiencies

Three BODIPY derivatives substituted at meso position by aryl (phenyl or N,N-dimethylaminonaphthyl) groups and functionalized at position 2 with electron-withdrawing formyl or electron deficient benzimidazole heterocycle were synthesized and completely characterized by the usual spectroscopic techniques. A comprehensive photophysical study showed a remarkable enhancement of the singlet oxygen sensitization quantum yield for meso-substituted dimethylaminonaphthyl BODIPY with the decrease of the dielectric constant of the solvent (0.02–0.04 for dimethylsulfoxide vs. 0.84 in toluene). In toluene the triplet state formation for the aminonaphthyl substituted BODIPY's was found to be mediated by the intramolecular charge transfer state (ICT), whereas in polar solvents triplet state formation is hindered by fast recombination of the ICT state. Pump-probe transient absorption spectroscopy was used to characterize the photoinduced dynamics of the BODIPY derivatives from the femtosecond to the nanosecond time scale.

Group IIA alkaline-earth metal phthalocyanines: Synthesis, photoluminescence and singlet oxygen formation

We have synthesized Group IIA alkaline-earth metal phthalocyanine (PC) complexes (BePC, MgPC, CaPC, SrPC, BaPC) using four different methods to study their photoluminescence and singlet oxygen photosensitizing properties. Their UV–Vis electronic absorption, fluorescence emission and excitation spectra, fluorescence quantum yields and lifetimes, as well as singlet oxygen formation quantum yields have been measured. The results reveal that alkali-earth metal PCs (BePC, MgPC, CaPC, SrPC, and BaPC) have high fluorescence quantum yields (0.50 to 0.70) and long fluorescence lifetimes (7.1 to 7.9 ns). SrPC, and BaPC also show high quantum yield of singlet oxygen formation (0.40 to 0.50). Therefore, alkaline-earth metal phthalocyanines are appealing fluorescent sensors and singlet oxygen photosensitizers for photodynamic therapy of tumors.

Heavy-Atom Free spiro Organoboron Complexes As Triplet Excited States Photosensitizers for Singlet Oxygen Activation.

Herein, we present a new strategy for the development of efficient heavy-atom free singlet oxygen photosensitizers based on rigid borafluorene scaffolds. Physicochemical properties of borafluorene complexes can be easily tuned through the choice of ligand, thus allowing exploration of numerous organoboron structures as potent 1O2 sensitizers. The singlet oxygen generation quantum yields of studied complexes vary in the range of 0.55–0.78. Theoretical calculations reveal that the introduction of the borafluorene moiety is crucial for the stabilization of a singlet charge transfer state, while intersystem crossing to a local triplet state is facilitated by orthogonal donor–acceptor molecular architecture. Our study shows that quantitative oxidation of selected organic substrates can be achieved in 20–120 min of irradiation with only 0.05 mol % loading of a photocatalyst.

Singlet oxygen‐responsive photorelease of tyramine

AbstractA system capable of photoreleasing tyramine has been developed. The photodonor system consists of an isoindole cage of tyramine and a biscyclometalated Ir(III) complex singlet oxygen (1O2) photosensitizer. Photoirradiation at a wavelength of 365 nm produces tyramine, as evidenced by mass spectrometry and 1H NMR spectroscopy. The photorelease proceeds through two steps involving the formation of a Diels–Alder‐type [2 + 4] cycloadduct of 1O2, followed by the slow and spontaneous decomposition of the adduct into tyramine and 2‐benzoylbenzophenon.

Singlet Oxygen Photosensitization Using Graphene-Based Structures and Immobilized Dyes: A Review

Singlet Oxygen Photosensitization Using Graphene-Based Structures and Immobilized Dyes: A Review

Estimation of Singlet Oxygen Quantum Yield Using Novel Green-Absorbing Baird-type Aromatic Photosensitizers.

We report two new organic green-absorbing singlet oxygen (1 O2 ) photosensitizers: Quinoidal naphthyl thioamide (QDM) and bis-iodol-dipyrrolonaphthyridine-dione (I2 -DPND), with triplet energies of 40.8 and 47.5 kcal mol-1 (at 77 K in a glassy matrix) , respectively. The UV-vis absorption and emission characteristics of QDM and I2 -DPND are similar to other commercially available organic 1 O2 photosensitizers such as Rose Bengal, which was used as standard/reference to estimate the 1 O2 quantum yield (Φ∆ ) of the chromophores under study. Using 9,10-diphenylanthracene (DPA) as an 1 O2 quencher, we estimated the Φ∆ ≈ 67-85% for QDM and Φ∆ ≈ 25-32% for I2 -DPND. The discrepancy in the Φ∆ values could be explained by the apparent photo-decomposition of the later dye. Nevertheless, the high Φ∆ value for QDM is unprecedented, as this chromophore exhibits relatively low structural complexity and could further be derivatized to create novel photodynamic agents.

The Polarity of Fullerene C60 Derivatives for Enhanced Photodynamic Inactivation.

In this article, four novel fulleropyrrolidines derivatives were synthesized to study how the effect of polarity and positive charge distribution can influence the efficacy of photodynamic inactivation treatments to kill bacteria. The design of the photosensitizers was based on DFT calculations that allowed us to estimate the dipolar moment of the molecules. Neutral compounds bearing N-methyl bis-acetoxy-ethyl (1) and bis-hydroxyethyl (2) amine were the starting material to obtain the dicationic analogs N,N-dimethyl bis-methoxyethyl (3), and bis-acetoxy-ethyl) (4) methylammonio. As expected from fullerene C60 derivatives, compounds 1-4 absorb in the UV region, with a peak at 430 nm, a broader range of absorption up to 710 nm, and exhibit weak fluorescence emission in toluene and reverse micelles. In the biomimetic AOT micellar system, the highest singlet oxygen photosensitization was found for compounds 1, followed by 3, 2, and 4. Whereas 4 was the most effective reducing nitro blue tetrazolium in the presence of β-NADH. The influence of type I and type II mechanism on the photodynamic activity of compounds 3 and 4 was further examined in the presence of L-tryptophan and two reactive oxygen species scavengers. In vitro experiments indicated that the compounds with the highest dipolar moments, 3 (37.19 D) and 4 (38.46 D), inactivated methicillin-resistant Staphylococcus aureus and Escherichia coli bacteria using an energy dose <2.4 J cm-2 . No inactivation was observed for the neutral analogs with the lowest dipolar moments. These findings help to optimize sensitizer structures to improve photodynamic inactivation.

Photodynamic Inactivation of Pseudomonas aeruginosa by PHEMA Films Loaded with Rose Bengal: Potentiation Effect of Potassium Iodide.

Four formulations have been used to produce different poly(2-hydroxyethyl methacrylate) (PHEMA) thin films, containing singlet oxygen photosensitizer Rose Bengal (RB). The polymers have been characterized employing Thermogravimetric Analysis (TGA), Attenuated Total Reflectance Fourier Transform Infrared Spectroscopy (ATR-FTIR) and UV-vis Absorption Spectroscopy. When irradiated with white light (400–700 nm) films generated singlet oxygen (1O2), as demonstrated by the reactivity with 1O2 trap 9,10-dimethylanthracene (DMA). Material with the highest RB loading (polymer A4, 835 nmol RB/g polymer) was able to perform up to ten cycles of DMA oxygenation reactions at high conversion rates (ca. 90%). Polymer A4 was also able to produce the complete eradication of a Pseudomonas aeruginosa planktonic suspension of 8 log10 CFU/mL, when irradiated with white light (total dose 72 J/cm2). The antimicrobial photodynamic effect was remarkably enhanced by adding potassium iodide (100 mM). In such conditions the complete bacterial reduction occurred with a total light dose of 24 J/cm2. Triiodide anion (I3−) generation was confirmed by UV-vis absorption spectroscopy. This species was detected inside the PHEMA films after irradiation and at concentrations ca. 1 M. The generation of this species and its retention in the matrix imparts long-lasting bactericidal effects to the RB@PHEMA polymeric hydrogels. The polymers here described could find potential applications in the medical context, when optimized for their use in everyday objects, helping to prevent bacterial contagion by contact with surfaces.

Understanding delayed fluorescence and triplet decays of Protoporphyrin IX under hypoxic conditions.

Photosensitizers of singlet oxygen exhibit three main types of reverse intersystem-crossing (RISC): thermally activated, triplet-triplet annihilation, and singlet oxygen feedback. RISC can be followed by delayed fluorescence (DF) emission, which can provide important information about the excited state dynamics in the studied system. An excellent model example is a widely used clinical photosensitizer Protoporphyrin IX, which manifests all three mentioned types of RISC and DF. Here, we estimated rate constants of individual RISC and DF processes in Protoporphyrin IX in dimethylformamide, and we showed how these affect triplet decays and DF signals under diverse experimental conditions, such as a varying oxygen concentration or excitation intensity. This provided a basis for a general discussion on guidelines for a more precise analysis of long-lived signals. Furthermore, it has been found that PpIX photoproducts and potential transient excited complexes introduce a new overlapping delayed luminescence spectral band with a distinct lifetime. These findings are important for design of more accurate biological oxygen sensors and assays based on DF and triplet lifetime.

Characterization of Nannochloropsis oceanica Rose Bengal Mutants Sheds Light on Acclimation Mechanisms to High Light When Grown in Low Temperature.

A barrier to realizing Nannochloropsis oceanica's potential for omega-3 eicosapentaenoic acid (EPA) production is the disparity between conditions that are optimal for growth and those that are optimal for EPA biomass content. A case in point is temperature: higher content of polyunsaturated fatty acid, and especially EPA, is observed in low-temperature (LT) environments, where growth rates are often inhibited. We hypothesized that mutant strains of N. oceanica resistant to the singlet-oxygen photosensitizer Rose Bengal (RB) would withstand the oxidative stress conditions that prevail in the combined stressful environment of high light (HL; 250 μmol photons m-2 s-1) and LT (18°C). This growth environment caused the wild-type (WT) strain to experience a spike in lipid peroxidation and an inability to proliferate, whereas growth and homeostatic reactive oxygen species levels were observed in the mutant strains. We suggest that the mutant strains' success in this environment can be attributed to their truncated photosystem II antennas and their increased ability to diffuse energy in those antennas as heat (non-photosynthetic quenching). As a result, the mutant strains produced upward of four times more EPA than the WT strain in this HL-LT environment. The major plastidial lipid monogalactosyldiacylglycerol was a likely target for oxidative damage, contributing to the photosynthetic inhibition of the WT strain. A mutation in the NO10G01010.1 gene, causing a subunit of the 2-oxoisovalerate dehydrogenase E1 protein to become non-functional, was determined to be the likely source of tolerance in the RB113 mutant strain.

A nanocellulose-based platform towards targeted chemo-photodynamic/photothermal cancer therapy

Cellulose nanocrystals (CNCs) have advantages as drug delivery carriers because of their biocompatibility and the presence of hydroxyl groups which favor chemical modification and drug binding. The present study describes the development of novel multifunctional rod-like CNCs-based carriers as therapeutic platforms: CNCs were hybridized with folic acid for actively targeting tumor cells, carbon dots (Cdots) for both imaging and photodynamic/photothermal treatments and doxorubicin (DOX) as an anticancer drug. Hybridized carriers displayed excellent drug-loading capacity. Moreover, Cdots-containing hybrids showed fluorescence and photosensitized singlet oxygen generation and photothermal behavior. Carriers exhibited pH-sensitive drug release because of changing interactions with DOX, and this release proved to be effective against in vitro cervical cancer cells, as evidenced by dose-dependent reduced cellular viabilities. Additionally, DOX release was promoted by light irradiation and the photodynamic behavior by reactive oxygen species was confirmed. These results demonstrate the potential of multifunctional CNCs-based carriers as platforms for multimodal photodynamic/photothermal-chemotherapy.

Porphyrin-Containing MOFs and COFs as Heterogeneous Photosensitizers for Singlet Oxygen-Based Antimicrobial Nanodevices.

Visible-light irradiation of porphyrin and metalloporphyrin dyes in the presence of molecular oxygen can result in the photocatalytic generation of singlet oxygen (1O2). This type II reactive oxygen species (ROS) finds many applications where the dye, also called the photosensitizer, is dissolved (i.e., homogeneous phase) along with the substrate to be oxidized. In contrast, metal-organic frameworks (MOFs) are insoluble (or will disassemble) when placed in a solvent. When stable as a suspension, MOFs adsorb a large amount of O2 and photocatalytically generate 1O2 in a heterogeneous process efficiently. Considering the immense surface area and great capacity for gas adsorption of MOFs, they seem ideal candidates for this application. Very recently, covalent-organic frameworks (COFs), variants where reticulation relies on covalent rather than coordination bonds, have emerged as efficient photosensitizers. This comprehensive mini review describes recent developments in the use of porphyrin-based or porphyrin-containing MOFs and COFs, including nanosized versions, as heterogeneous photosensitizers of singlet oxygen toward antimicrobial applications.

Novel group 14 element-bridged bithiophene dimers appended with terminal electron-withdrawing groups: Red-to-near infrared fluorescence and efficient photosensitized singlet oxygen generation

A series of group 14 element bridged bithiophene dimers bearing dicyanovinyl (DC) or thiobarbiturylmethylene (TB) electron-withdrawing groups at the molecular termini (series E-1a and series E-1b; E = C, Si, and Ge) were developed as red absorbing singlet oxygen (1O2) photosensitizing dyes. These dyes exhibited broad and strong absorption in the red-to-deep red regions and efficient red-to-NIR fluorescence (photoluminescence quantum yield ΦPL; 0.32–0.45 in dichloromethane). The terminal TB groups led to more red-shifted electronic absorption and fluorescence emission than the DC groups, owing to more intense intramolecular charge transfer. The 1O2 generation photosensitization abilities of series E-1a and series E-1b were evaluated by the 1,3-diphenylisobenzofuran method, and large 1O2 generation quantum yields (ΦΔ) were obtained upon irradiation of 630 nm light, ranging from 0.35 to 0.68 in dichloromethane. Therefore, these dyes show fluorescence properties as well as 1O2 generation photosensitization ability. Introduction of bromo groups at the 3,3′-positions in Si-1a and Si-1b afforded the new potential photosensitizers Si-2a and Si-2b that accompanied blue-shifted electronic absorption as well as blue-shifted fluorescence emission with the low ΦPL. Unfortunately, as Si-2a exhibited quite low absorptivity in the red region, 1O2 generation photosensitization with 630 nm light was not evaluated for Si-2a. On the other hand, Si-2b showed significantly efficient 1O2 generation photosensitization upon irradiation of 630 nm light (ΦΔ; 0.88). Considering the result of theoretical calculations showing that the bromo groups effectively contribute to the frontier orbitals, these bromo substituents should facilitate the intersystem crossing from the excited singlet state to the triplet excited state through the strong spin‒orbit coupling.

Understanding and Controlling the Efficiency of Au24 m(SR)18 Nanoclusters As Singlet-Oxygen Photosensitizers

Singlet oxygen, 1O2, is an excited oxygen species [1] that can be efficiently generated by photosensitizers. These are molecules that upon photoexcitation eventually generate a triplet excited state that can be quenched by the ground state oxygen triplet, 3O2, to form 1O2 [1]. The latter finds uses in many applications, especially organic synthesis, photocatalysis, and photodynamic therapy [2-4]. On the other hand, its possible use depends on its effective lifetime, as 1O2 is a very reactive species that decays via quenching by the solvent and/or interactions with other molecules in solution [1]. This is the case of the photosensitizers themselves, and this may result in a serious drawback for efficient applications of 1O2. In this area, the use of monolayer-protected metal nanoclusters as photosensitizers is limited. We carried out a systematic study on the photosensitizer behavior of a series of molecular Au24M(SR)18 clusters [5,6], where the R group and the doping metal M were varied. Electrochemical characterization allowed us to select the best R/M combinations to test a sufficiently large oxidation-potential range. 1O2 detection was carried out by time-resolved electron paramagnetic resonance [7], which allowed us to determine 1O2 lifetimes spanning more than one order of magnitude (on the microsecond time scale), depending on the specific cluster employed. Among the main results, we found that: (i) a more positive potential for the oxidation of the cluster is matched by a (beneficial) longer 1O2 lifetime; (ii) proper design of the electrochemical properties of the cluster yields the same results as those of reference photosensitizers, despite the unprecedented low oxidation potentials exhibited by these clusters; (iii) kinetic analysis based on the Marcus theory provides important insights into the mechanism of the quenching of 1O2 by gold nanoclusters, how to control it, and why properly doped gold nanoclusters may perform very well in 1O2 photosensitization [8]. Further effects, detected by electrochemistry, are under investigation.

Broad-Spectrum Photo-Antimicrobial Polymers Based on Cationic Polystyrene and Rose Bengal.

New strategies to fight bacteria and fungi are necessary in view of the problem of iatrogenic and nosocomial infections combined with the growing threat of increased antimicrobial resistance. Recently, our group has prepared and described two new readily available materials based on the combination of Rose Bengal (singlet oxygen photosensitizer) and commercially available cationic polystyrene (macroporous resin Amberlite® IRA 900 or gel-type resin IRA 400). These materials showed high efficacy in the antimicrobial photodynamic inactivation (aPDI) of Pseudomonas aeruginosa. Here, we present the photobactericidal effect of these polymers against an extended group of pathogens like Escherichia coli, Enterococcus faecalis, Staphylococcus aureus, and the opportunistic yeast Candida albicans using green light. The most interesting finding is that the studied materials are able to reduce the population of both Gram-positive and Gram-negative bacteria with good activity, although, for C. albicans, in a moderate manner. In view of the results achieved and especially considering the inexpensiveness of these two types of photoactive polymers, we believe that they could be used as the starting point for the development of coatings for self-disinfecting surfaces.

Stable Transfection of the Singlet Oxygen Photosensitizing Protein SOPP3: Examining Aspects of Intracellular Behavior†

Protein-encased chromophores that photosensitize the production of reactive oxygen species, ROS, have been the center of recent activity in studies of oxidative stress. One potential attribute of such systems is that the local environment surrounding the chromophore, and that determines the chromophore's photophysics, ideally remains constant and independent of the global environment into which the system is placed. Therefore, a protein-encased sensitizer localized in the mitochondria would arguably have the same photophysics as that protein-encased sensitizer at the plasma membrane, for example. One thus obtains a useful tool to study processes modulated by spatially localized ROS. One ROS of interest is singlet oxygen, O2 (a1 Δg ). We recently developed a singlet oxygen photosensitizing protein, SOPP, in which flavin mononucleotide, FMN, is encased in a re-engineered light-oxygen-voltage protein. One goal was to ascertain how a version of this system, SOPP3, which selectively makes O2 (a1 Δg ), invitro, behaves in a cell. We now demonstrate that SOPP3 undergoes exacerbated irradiation-mediated bleaching when expressed at either the plasma membrane or mitochondria in stable cell lines. We find that the environment around the SOPP3 system affects the bleaching rate, which argues against one of the key suppositions in support of a protein-encased chromophore.

Metal-Enhanced Photosensitization of Singlet Oxygen (ME1O2) from Brominated Carbon Nanodots on Silver Nanoparticle Substrates

The deleterious effects of reactive oxygen species (ROS), including singlet oxygen (1O2), on biological systems have cultivated widespread interest in fields ranging from therapeutic techniques to sterilization materials. Researchers have, for example, sought to capitalize on the oxidative damage from singlet oxygen to treat tumors as well as to kill antibiotic resistant bacteria. To generate 1O2 in a controllable manner, photosensitizers are optimized to generate 1O2 from ground state oxygen (3O2) when excited by light. When considering applications of photosensitization, favorable properties include high 1O2 yield, low synthetic complexity, and minimal cost. Previously, studies have shown that plasmonic nanoparticles are able to amplify the photosensitization of 1O2 from small molecule photosensitizers in a mechanism similar to metal-enhanced fluorescence (MEF), thereby improving yield. A recent study from our lab has demonstrated that brominated carbon nanodots, which are an inexpensive and simple-to-collect as a hydrocarbon combustion byproduct, generate reactive oxygen species that can be used for antimicrobial photodynamic inactivation of bacteria. Herein we investigate the combination of these advantageous properties. Using the turn-on fluorescent probe Singlet Oxygen Sensor Green™ to detect 1O2, we report the metal-enhanced photosensitization of 1O2 by brominated dots in silvered Quanta Plate™ wells. These results provide a promising direction for the potential optimization of carbon nanodot-based agents in light-activated antimicrobial materials.

Comparative singlet oxygen photosensitizer efficiency of berberine, rose bengal, and methylene blue by time course nuclear magnetic resonance (NMR) monitoring of a photochemical 4+2 cycloaddition endoperoxide formation

Berberine, a natural product alkaloid, has been shown to exert biological activity via in situ production of singlet oxygen when photo irradiated. Berberine utilizes singlet oxygen in its putative mechanism of action, wherein it forms an activated complex with DNA and photosensitizes triplet oxygen to singlet oxygen to specifically oxidize guanine residues, thereby halting cell replication and leading to cell death. This has potential application in photodynamic therapy, alongside other such compounds which also act as photosensitizers and produce singlet oxygen in situ. The quantification of singlet oxygen in various photosensitizers, including berberine, is essential for determining their photosensitizer efficiencies. We postulated that the singlet oxygen produced by photoirradiation of berberine would be superior in terms of singlet oxygen production to the aforementioned photosensitizers when irradiated with UV light, but inferior under visible light conditions, due to its strong absorbance of UV wavelengths. Here, we report the usage of time course 1H nuclear magnetic resonance (NMR) spectroscopy to trap singlet oxygen via a 4+2 cycloaddition with terpinene, as well as theoretical calculations by time-dependent density functional theory (TD-DFT) towards quantification of berberine's singlet oxygen production against two known photosensitizers, methylene blue and rose bengal, to determine berberine’s efficacy as a singlet oxygen photosensitizer. We envision that similar processes can be utilized for the evaluation of berberine analogs or other photosensitizing agents and the identification of other potential medicinally significant singlet oxygen photosensitizers.

Synthesis, characterization, and cellular investigations of porphyrin- and chlorin-indomethacin conjugates for photodynamic therapy of cancer.

Indomethacin is a potent non-steroidal anti-inflammatory drug (NSAID) with a strong selective inhibitor activity towards cyclooxygenase-2 (COX-2), an enzyme that is highly overexpressed in various tumour cells, being involved in tumourigenesis. Concomitantly, porphyrins have gained much attention as promising photosensitizers (PSs) for the non-invasive photodynamic therapy (PDT) of cancer. Herein, we report the design, and determine the singlet oxygen generation capacity and in vitro cellular toxicity of porphyrin- and chlorin-indomethacin conjugates (P2-Ind and C2-Ind). Both the conjugates were obtained in high yields and were characterized by 1H, 19F and 13C NMR as well as by high resolution mass spectrometry. The singlet oxygen generation properties were assessed by the 1,3-diphenylisobenzofuran singlet oxygen trap method, which showed that C2 and C2-Ind are the best singlet oxygen photosensitizers. In addition, it was found that the presence of indomethacin did not influence the singlet oxygen generation of porphyrin or chlorin. Cytotoxicity studies of the conjugate in human HEp2 cells revealed that the porphyrin- and chlorin-indomethacin conjugates have similar dark cytotoxicities, while chlorin C2 was shown to be the most phototoxic. Despite having lower cellular uptake than C2-Ind after 24 hours, chlorin C2 had a broad localization in HEp2 cells while the chlorin-indomethacin conjugate C2-Ind could be detected in the form of small aggregates. DFT calculations were performed to shed light on the reaction energy involved in the formation of the indomethacin conjugates and to compare the relative stability of selected isomers in solution. Moreover, the calculated energy of their first excited triplet state structures confirmed their use as suitable photosensitizers to generate singlet oxygen for PDT.

From photosensitizers to light harvesters adapting the molecular structure in all-BODIPY assemblies.

Herein we detail a protocol to design dyads and triads based solely on BODIPY dyes as halogen-free singlet oxygen photosensitizers or energy transfer molecular cassettes. The conducted photonic characterization reveals the key role of the BODIPY-BODIPY linkage to finely modulate the balance between the triplet state population and fluorescence decay.