Time-resolved Singlet Oxygen Luminescence Research Articles

Monitoring of time-resolved singlet oxygen luminescence at 1270nm by an optical fiber detection system

Singlet oxygen (1O2) is the main cytotoxic substance in Type II photodynamic therapy (PDT). The luminescence of 1O2 at 1270[Formula: see text]nm is extremely weak with a low quantum yield, making the direct detection of 1O2 at 1270[Formula: see text]nm very challenging. In this study, a set of highly sensitive optical fiber detection system is built up to detect the luminescence of photosensitized 1O2. We use this system to test the luminescence characteristics of 1O2 in pig skin tissue ex vivo and mouse auricle skin in vivo. The experimental results show that the designed system can quantitatively detect photosensitized 1O2 luminescence. The 1O2 luminescence signal at 1270[Formula: see text]nm is successfully detected in pig skin ex vivo. Compared with RB in an aqueous solution, the lifetime of 1O2 increases to [Formula: see text]s in pig skin tissue ex vivo. Experiments on living mice suggest that an enhancement of 1O2 intensity with the increase of the TMPyP concentration. When the dose is 25[Formula: see text]mg/kg, the vasoconstriction can reach more than 80%. The results of this study hold the potential application for clinical PDT dose monitoring using an optical fiber detection system.

Enhanced Optics for Time-Resolved Singlet Oxygen Luminescence Detection

Singlet oxygen luminescence dosimetry (SOLD) is a highly promising direct monitoring method for photodynamic therapy (PDT) in the treatment of cancer. Early SOLD systems have been hampered by inefficient excitation, poor optical collection and immature infrared single photon detection technology. We report carefully engineered improvements addressing all of these deficiencies. We use a supercontinuum source with a tunable filter to precisely target the peak absorption wavelength of the chosen photosensitizer; we have designed a compact and versatile optical package for precise alignment; we have successfully employed state-of-the-art superconducting photon counting technologies. Through these improvements, we can achieve histogram acquisition from a photosensitizer in solution test sample. This setup opens the pathway to physiological SOLD studies for PDT dosimetry.

Photodynamic inactivation of Escherichia coli – Correlation of singlet oxygen kinetics and phototoxicity

Photodynamic inactivation (PDI) of bacteria may play a major role in facing the challenge of the ever expanding antibiotic resistances. Here we report about the direct correlation of singlet oxygen luminescence kinetics and phototoxicity in E. coli cell suspension under PDI using the widely applied cationic photosensitizer TMPyP. Through direct access to the microenvironment, the time resolved investigation of singlet oxygen luminescence plays a key role in understanding the photosensitization mechanism and inactivation pathway. Using the homemade set-up for highly sensitive time resolved singlet oxygen luminescence detection, we show that the cationic TMPyP is localized predominantly outside the bacterial cells but in their immediate vicinity prior to photodynamic inactivation. Throughout following light exposure, a clear change in singlet oxygen kinetics indicates a redistribution of photosensitizer molecules to at least one additional microenvironment. We found the signal kinetics mirrored in cell viability measurements of equally treated samples from same overnight cultures conducted in parallel: A significant drop in cell viability of the illuminated samples and stationary viability of dark controls. Thus, for the system investigated in this work – a Gram-negative model bacteria and a well-known PS for its PDI – singlet oxygen kinetics correlates with phototoxicity. This finding suggests that it is well possible to evaluate PDI efficiency directly via time resolved singlet oxygen detection.

Prospects of in vivo singlet oxygen luminescence monitoring: Kinetics at different locations on living mice

BackgroundSinglet oxygen observation is considered a valuable tool to assess and optimize PDT treatment. In complex systems, such as tumors in vivo, only the direct, time-resolved singlet oxygen luminescence detection can give reliable information about generation and interaction of singlet oxygen. Up to now, evaluation of kinetics was not possible due to insufficient signal-to-noise ratio. Here we present high signal-to-noise ratio singlet oxygen luminescence kinetics obtained in mouse tumor model under PDT relevant conditions. MethodsA highly optimized system based on a custom made laser diode excitation source and a high aperture multi-furcated fiber, utilizing a photomultiplier tube with a multi photon counting device was used. ResultsLuminescence kinetics with unsurpassed signal-to-noise ratio were gained from tumor bearing nude mice in vivo upon topic application, subcutaneous injection as well as intravenous injection of different photosensitizers (chlorin e6 and dendrimer formulations of chlorin e6). Singlet oxygen kinetics in appropriate model systems are discussed to facilitate the interpretation of complex kinetics obtained from in vivo tumor tissue. ConclusionsThis is the first study addressing the complexity of singlet oxygen luminescence kinetics in tumor tissue. At present, further investigations are needed to fully explain the processes involved. Nevertheless, the high signal-to-noise ratio proves the applicability of direct time-resolved singlet oxygen luminescence detection as a prospective tool for monitoring photodynamic therapy.

Time-resolved singlet-oxygen luminescence detection with an efficient and practical semiconductor single-photon detector.

In clinical applications, such as PhotoDynamic Therapy, direct singlet-oxygen detection through its luminescence in the near-infrared range (1270 nm) has been a challenging task due to its low emission probability and the lack of suitable single-photon detectors. Here, we propose a practical setup based on a negative-feedback avalanche diode detector that is a viable alternative to the current state-of-the art for different clinical scenarios, especially where geometric collection efficiency is limited (e.g. fiber-based systems, confocal microscopy, scanning systems etc.). The proposed setup is characterized with Rose Bengal as a standard photosensitizer and it is used to measure the singlet-oxygen quantum yield of a new set of photosensitizers for site-selective photodynamic therapy.

Luminescence investigation of photosensitizer distribution in skin: correlation of singlet oxygen kinetics with the microarchitecture of the epidermis

This is the first study showing that singlet oxygen kinetics of topically applied photosensitizers coincides with the microarchitecture of skin, e.g., fissures and hair follicles. The kinetics indicate a chemical interaction of singlet oxygen with the skin, which allows differentiating between residual crème, e.g., in the follicular orifice, and photosensitizer penetrated into the skin. We show the feasibility of an easy-to-use fiber optic application providing the opportunity for in situ investigation, as well as a setup with focused optics for high-resolution two-dimensional scanning of singlet oxygen luminescence kinetics in skin samples. The results show that time-resolved singlet oxygen luminescence detection in tissue is a desirable tool for medical therapy, diagnostics, and evaluation of singlet oxygen interaction with biological environments.

Highly sensitive time resolved singlet oxygen luminescence detection using LEDs as the excitation source

For the first time singlet oxygen luminescence kinetics in living cells were detected at high precision using LED light for excitation. As LED technology evolves, the light intensity emitted by standard LEDs allows photosensitized singlet oxygen luminescence detection in solution and cell suspensions. We present measurements superior to those of most actual laser powered setups regarding precision of singlet oxygen kinetics in solutions and cell suspensions. Data presented here show that LED based setups allow the determination of the photosensitizer triplet and singlet oxygen decay times in vitro with an accuracy of 0.1 μs. This enables monitoring of the photosensitizer efficiency and interaction with the cellular components using illumination doses small enough not to cause cell death.

Time-resolved singlet oxygen luminescence detection under photodynamic therapy relevant conditions: comparison ofex vivoapplication of two photosensitizer formulations

Singlet oxygen plays a crucial role in photo-dermatology and photodynamic therapy (PDT) of cancer. Its direct observation by measuring the phosphorescence at 1270 nm, however, is still challenging due to the very low emission probability. It is especially challenging for the time-resolved detection of singlet oxygen kinetics in vivo which is of special interest for biomedical applications. Photosensitized generation of singlet oxygen, in pig ear skin as model for human skin, is investigated here. Two photosensitizers (PS) were topically applied to the pig ear skin and examined in a comparative study, which include the amphiphilic pheophorbide-a and the highly hydrophobic perfluoroalkylated zinc phthalocyanine (F64PcZn). Fluorescence microscopy indicates the exclusive accumulation of pheophorbide-a in the stratum corneum, while F64PcZn can also accumulate in deeper layers of the epidermis of the pig ear skin. The kinetics obtained with phosphorescence measurements show the singlet oxygen interaction with the PS microenvironment. Different generation sites of singlet oxygen correlate with the luminescence kinetics. The results show that singlet oxygen luminescence detection can be used as a diagnostic tool, not only for research, but also during treatment. The detection methodology is suitable for the monitoring of chemical quenchers’ oxidation as well as saturation at singlet oxygen concentration levels relevant to PDT treatment protocols.

Direct Observation and Quantitative Characterization of Singlet Oxygen in Aqueous Solution upon UVA Excitation of 6-Thioguanines

The incorporation of 6-thioguanine (6-TG) into DNA increases the risk of (1)O(2)-initiated skin cancer. We herein provide the first report on quantitative characterization of the photoactivity of 6-thioguanines including 6-TG and 6-thioguanosine. Time-resolved singlet oxygen luminescence was observed directly for the first time after UVA irradiation of 6-thioguanines in both CHCN(3) and aqueous solutions. Their photosensitization was characterized by the quantum yield of singlet oxygen production, showing a dramatic decrease over time from the initial 0.49-0.58 to zero. Experiments performed on both 6-TG and 6-thioguanosine did not show any significant difference in the quantum yield of singlet oxygen production, indicating that there was no potential participation of 7H- and 9H-tautomers. Our findings provide a primary basis for a better understanding of molecular events of thiopurine drugs in biological systems.

New insights to primary photodynamic effects – Singlet oxygen kinetics in living cells

The kinetics of chemical singlet oxygen quencher consumption inside living cells during low dose illumination was revealed via time resolved singlet oxygen luminescence detection. Deviations of the measured data from the common theoretical model for 1O 2 kinetics forced the authors to consider a one-dimensional diffusion model for description of the kinetics of singlet oxygen generated by membrane localized photosensitizers. Our observations reconcile seemingly contradictory reports presenting different values for the efficiency of singlet oxygen interaction with cells.

Synthesis and solvent dependence of the photophysical properties of [60]fullerene–sugar conjugates

A method for the synthesis of optically pure C 60 derivatives containing one or two d-galactose or d-glucose units is described. It involves the synthesis of sugar–malonate derivatives followed by a cyclopropanation reaction with C 60. The solvent dependence of the photophysical properties of the methano[60]fullerene–sugar derivatives was studied using nanosecond laser flash photolysis coupled with kinetic UV–vis absorption spectroscopy and time-resolved singlet oxygen luminescence measurements. The triplet properties of these fullerenes, including transient absorption spectra, molar absorption coefficients and quantum yield for the photosensitised production of 1O 2 were determined in toluene, benzonitrile and acetonitrile solutions. The transient absorption spectral profiles are solvent independent although small differences are observed in the transient absorption maximum: 720±5 nm for toluene, 710±5 nm for benzonitrile and 700±5 nm for acetonitrile. Triplet state molar absorption coefficients ( ε T) of C 60 derivatives vary from 9456±2090 M −1 cm −1, for compound 10 in toluene, and 15,272±4462 M −1 cm −1, for compound 6 in acetonitrile. Triplet state lifetimes ( τ T) for methano[60]fullerene–sugar derivatives, under our experimental conditions, are similar in toluene or benzonitrile solutions (47.5±1.1 μs≤ τ T≤51.4±2.0 μs) but are lower in acetonitrile solutions (31.8±0.6 μs≤ τ T≤43.0±1.1 μs). Toluene and benzonitrile solutions of C 60 derivatives have Φ Δ close to unity.