Measurements Of Singlet Oxygen Luminescence Research Articles

High Sensitivity Singlet Oxygen Luminescence Sensor Using Computational Spectroscopy and Solid-State Detector.

This paper presents a technique for high sensitivity measurement of singlet oxygen luminescence generated during photodynamic therapy (PDT) and ultraviolet (UV) irradiation on skin. The high measurement sensitivity is achieved by using a computational spectroscopy (CS) approach that provides improved photon detection efficiency compared to spectral filtering methodology. A solid-state InGaAs photodiode is used as the CS detector, which significantly reduces system cost and improves robustness compared to photomultiplier tubes. The spectral resolution enables high-accuracy determination and subtraction of photosensitizer fluorescence baseline without the need for time-gating. This allows for high sensitivity detection of singlet oxygen luminescence emission generated by continuous wave light sources, such as solar simulator sources and those commonly used in PDT clinics. The value of the technology is demonstrated during in vivo and ex vivo experiments that show the correlation of measured singlet oxygen with PDT treatment efficacy and the illumination intensity on the skin. These results demonstrate the potential use of the technology as a dosimeter to guide PDT treatment and as an analytical tool supporting the development of improved sunscreen products for skin cancer prevention.

Photodynamic Inhibition of Biofilm Forming Microorganisms on Surfaces

Microorganisms play an important role in our everyday life and in an ecological context. While a great number of species provide a benefit to humans, some of them can be terribly harmful due to the diseases they trigger. To remove or hinder those unwanted microorganisms has been a goal since their discovery. There are classical approaches to accomplish that, but the adaptability of microorganisms – especially the alarming development of antibiotic resistant strains - always calls for new methods. Once microorganisms grow in biofilms they are hard to come by and inadvertently spread infections are a fatal risk for immunodeficient people. Photodynamic Inactivation (PDI) has good potential as antimicrobial agent, which avoids most of aforementioned problems. Singlet oxygen is a highly reactive molecule against which there has been no development of countermeasures in biological systems besides provision of quenching molecules inside the cells. Therefore there is also little to no risk of resistance development when using PDI against microorganisms. Easy availability of light and oxygen in many of the potential target sites are just some advantages of PDI. Even though much less often associated with maladies, fungi can be similarly harmful. Due to their asexual life cycle and sporulation they are more robust against changes in environmental conditions. Their high robustness requires highly toxic biocides which in turn come with the risk of damaging the environment. Even seemingly, harmless microorganisms like algae and cyanobacteria show increasing tendencies to harm humans directly or indirectly and in symbiosis with fungi, they form even more resistant lichen, which pose new challenges concerning their removal. Biofilms on outdoor walls cause aesthetic problems and economic damage. Therefore, wall colors are often mixed with conventional biocides. However, biocides are increasingly designed towards higher efficacy while remaining environmentally persistent. This combination implies serious toxicity problems for all living organisms and whole ecosystems. Thus, an eco-friendly and non-harmful to human health alternative to conventional biocides in wall color is strongly recommended. The well-known photosensitizer TMPyP was used as an additive in commercially available facade paint. The generation of singlet molecular oxygen was shown using time resolved 2D measurements of the singlet oxygen luminescence. The photodynamic activity of the photosensitizer in the facade paint was demonstrated by phototoxicity tests with defined mold fungi and a mixture of microorganisms harvested from native outdoor biofilms as model organisms. Phototrophic biofilms are an important factor in biofouling and biodeterioration of building materials. The aim of this study was to examine the potential of PDI through generation of singlet oxygen as a possible alternative to biocides preventing the growth and formation of biofilms of these phototrophic organisms. Four photosensitizers were investigated according to their photoinhibitive effect on two strains of green algae using visible light for photoexcitation. The two anionic photosensitizers showed no significant phototoxicity whereas the two cationic photosensitizers caused a drastic reduction of biomass on a short time scale and also displayed long term inhibition of algae growth. Furthermore, to investigate the intrinsic mechanisms of PDI we studied the singlet oxygen luminescence kinetics - a method that proved to be a very effective approach towards understanding mechanisms on a cellular level. Here we present the first two-dimensional measurement of singlet oxygen kinetics in phototrophic microorganisms on surfaces during PDI. We established a system of reproducible algae samples on surfaces, incubated with two different cationic, antimicrobial potent photosensitizers. Fluorescence microscopy images indicate that one photosensitizer localizes inside the green algae while the other accumulates along the outer algae cell wall. A newly developed setup allows for the measurement of singlet oxygen luminescence on the green algae sample surfaces over several days. The kinetics of the singlet oxygen luminescence of both photosensitizers show different developments and a distinct change over time, corresponding with the differences in their localization as well as their photosensitization potential. While the complexity of the signal reveals a challenge for the future, this study incontrovertibly marks a crucial, inevitable step in the investigation of photodynamic inactivation of biofilms: it shows the feasibility of using the singlet oxygen luminescence kinetics to investigate photodynamic effects on surfaces and thus opens a field for numerous investigations. Figure 1

Development of Singlet Oxygen Luminescence Kinetics during the Photodynamic Inactivation of Green Algae.

Recent studies show the feasibility of photodynamic inactivation of green algae as a vital step towards an effective photodynamic suppression of biofilms by using functionalized surfaces. The investigation of the intrinsic mechanisms of photodynamic inactivation in green algae represents the next step in order to determine optimization parameters. The observation of singlet oxygen luminescence kinetics proved to be a very effective approach towards understanding mechanisms on a cellular level. In this study, the first two-dimensional measurement of singlet oxygen kinetics in phototrophic microorganisms on surfaces during photodynamic inactivation is presented. We established a system of reproducible algae samples on surfaces, incubated with two different cationic, antimicrobial potent photosensitizers. Fluorescence microscopy images indicate that one photosensitizer localizes inside the green algae while the other accumulates along the outer algae cell wall. A newly developed setup allows for the measurement of singlet oxygen luminescence on the green algae sample surfaces over several days. The kinetics of the singlet oxygen luminescence of both photosensitizers show different developments and a distinct change over time, corresponding with the differences in their localization as well as their photosensitization potential. While the complexity of the signal reveals a challenge for the future, this study incontrovertibly marks a crucial, inevitable step in the investigation of photodynamic inactivation of biofilms: it shows the feasibility of using the singlet oxygen luminescence kinetics to investigate photodynamic effects on surfaces and thus opens a field for numerous investigations.

Blue light kills Aggregatibacter actinomycetemcomitans due to its endogenous photosensitizers

The aim of this study was to demonstrate that the periodontal pathogen Aggregatibacter actinomycetemcomitans (AA) can be killed by irradiation with blue light derived from a LED light-curing unit due to its endogenous photosensitizers. Planktonic cultures of AA and Escherichia coli were irradiated with blue light from a bluephase® C8 light-curing unit with an emission peak at 460 nm, which is usually applied for polymerization of dental resins. A CFU-assay was performed for the analysis of viable bacteria after treatment. Moreover, bacterial cells were lysed and the lysed AA and E. coli were investigated for generation of singlet oxygen. Spectroscopic measurements of lysed AA and E. coli were performed and analyzed for characteristic absorption and emission peaks. A light dose of 150 J/cm(2) induced a reduction of ≥5 log10 steps of viable AA, whereas no effect of blue light was found against E. coli. Spectrally resolved measurements of singlet oxygen luminescence showed clearly that a singlet oxygen signal is generated from lysed AA upon excitation at 460 nm. Spectroscopic measurements of lysed AA exhibited characteristic absorption and emission peaks similar to those of known porphyrins and flavins. AA can be inactivated by irradiation with blue light only, without application of an exogenous photosensitizer. These results encourage further studies on the potential use of these blue light-mediated auto-photosensitization processes in the treatment of periodontitis for the successful inactivation of Aggregatibacter actinomycetemcomitans.

Photo-oxidation by singlet oxygen generated on nanoporous silicon in a LED-powered reactor

An annular flow photochemical reactor illuminated by UV and green (524 nm) light emitting diodes (LEDs) was characterised by a chemical actinometer. Very high efficiency of absorption of photons, most likely promoted by the specific orientation of LED elements in the reactor, was calculated based on the measured actinometry results. Generation of singlet oxygen mediated by nanoporous silicon under illumination by Ar+ laser, UV and green LEDs was demonstrated by indirect measurement of suppression of porous Si photoluminescence, and by direct measurements of singlet oxygen luminescence. The efficiency of reactor in singlet oxygen mediated reactions was tested using reaction of decomposition of diphenylbenzofuran. Estimated quantum yield of chemical reaction is about 34%.

One‐ and Two‐Photon Singlet Oxygen Generation with New Fluorene‐Based Photosensitizers

The spectral properties of new fluorene-based photosensitizers for efficient singlet oxygen production are investigated at room temperature and 77 K. Two-photon absorption (2PA) cross-sections of the fluorene derivatives are measured by the open aperture Z-scan method. The quantum yields of singlet oxygen generation under one- and two-photon excitation (phi(delta) and 2PAphi(delta), respectively), are determined by the direct measurement of singlet oxygen luminescence at approximately 1270 nm. The values of phi(delta) are independent of excitation wavelength, ranging from 0.6-0.9. The singlet oxygen quantum yields under two-photon excitation are 2PAphi(delta) approximately 1/2 phi(delta), indicating that the two processes exhibit the same mechanism of singlet oxygen production, independent of the mechanism of photon absorption.

Comparison of Singlet Oxygen Generation Efficiency between Water‐Soluble C60‐Diphenylaminofluorene Conjugates and Molecular Micelle‐like FC4S

Amphiphilic C60‐diphenylaminofluorene conjugates C60 (>DPAF‐EGn) exhibit good water‐solubility and form self‐assembled bilayer vesicles in dilute aqueous solution. Their singlet oxygen generation efficiency in DMF or H2O was evaluated by using time‐resolved measurements of singlet oxygen luminescence at 1270 nm upon photoexcitation by nanosecond pulse laser operated at 523 nm. By the application of different bandpass filters to cut off certain emission at wavelength ranges outside 1270 nm, a distinguishable peak in medium to high integrated luminescence intensity was collected at wavelengths between 1240 and 1300 nm. It provided confirmation and close correlation of emitted photon counts measured for these fullerene derivatives to singlet oxygen. In contrast to apparently high singlet oxygen generation efficiency of FC4S in H2O, an observed low singlet oxygen production rate from aqueous solutions of C60 (>DPAF‐EG14) and C60 (>DPAF‐EG45) was interpreted as the result of the occurrence of competitive ultrafast intramolecular electron transfer going from DPAF moiety to C60 cage moiety. That eliminated largely the possibility of energy transfer process for fullerenyl intersystem crossing to generate 3C*60(>DPAF‐EGn). Instead, radical ion‐pairs containing molecules C60 −· [>(DPAF‐EGn)+·] were produced via the intramolecular electron transfer process in the photoexcited transient state.

Determination of the electron transfer parameters of a covalently linked porphyrin-quinone with mesogenic substituents — optical spectroscopic studies in solution

The photoinduced electron transfer (PET) of a covalently linked porphyrin-quinone with mesogenic substituents was studied using visible and near-IR (NIR) spectroscopy. Mesogenic substituents were introduced at the porphyrin moiety in order to mimic the anisotropic membrane properties of the native reaction centre of photosynthesis. Photophysical characterization of this system in homogeneous solution is a prerequisite for a better understanding of the effects occurring in anisotropic medium. For this reason, we studied the fluorescence and phosphorescence quenching and lifetime of the charge-separated state. Time-resolved fluorescence measurements indicated an effective singlet PET. The complete set of PET parameters was calculated using Marcus theory of non-adiabatic electron transfer (ET). Steady state measurement of singlet oxygen luminescence, which allows indirect access to phosphorescence quenching, indicated that no triplet PET was involved in the decay processes. Using transient absorption spectroscopy, the lifetime of the charge-separated state was found to be 1.9 ns.