Photochemistry and Photobiology

Cover image for Vol. 90 Issue 4

Edited By: Jean Cadet

Impact Factor: 2.684

ISI Journal Citation Reports © Ranking: 2013: 36/74 (Biophysics); 161/291 (Biochemistry & Molecular Biology)

Online ISSN: 1751-1097

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  • RESEARCH ARTICLE: The Number of Accumulated Photons and the Quality of Stimulated Emission Depletion Lifetime Images

    RESEARCH ARTICLE: The Number of Accumulated Photons and the Quality of Stimulated Emission Depletion Lifetime Images

    Time binning is used to increase the number of photon counts in the peak channel of stimulated emission depletion fluorescence lifetime decay curves to determine how it affects the resulting lifetime image. The fluorescence lifetime of the fluorophore, Alexa Fluor 594 phalloidin, bound to F-actin is probed in cultured S2 cells at a spatial resolution of ~40 nm. This corresponds to a 10-fold smaller probe volume compared to confocal imaging, and a reduced number of photons contributing to the signal. Pixel-by-pixel fluorescence lifetime measurements and error analysis show that an average of 40 ± 30 photon counts in the peak channel with a signal-to-noise ratio of 20 is enough to calculate a reliable fluorescence lifetime from a single exponential fluorescence decay. No heterogeneity in the actin cytoskeleton in different regions of the cultured cells was measured in the 40–400 nm spatial regime.

  • RESEARCH NOTE: Preparation and Characterization of Uniform Near IR Polystyrene Nanoparticles

    RESEARCH NOTE: Preparation and Characterization of Uniform Near IR Polystyrene Nanoparticles

    Biomaterials for in vivo fluorescence imaging are required to be biocompatible, nontoxic, photostable and highly fluorescent. Fluorescence must be in the near infrared (NIR) region of the electromagnetic spectrum to avoid absorption and autofluorescence of endogenous tissues. NIR fluorescent polystyrene nanoparticles may be considered ideal biomaterials for in vivo imaging applications. These NIR nanoparticles were prepared by a swelling process of polystyrene template nanoparticles with a hydrophobic NIR dye dissolved in a water-miscible swelling solvent, a method developed for preparation of nonbiodegradable nanoparticles, for NIR fluorescent bioimaging applications. This method overcomes common problems that occur with dye entrapment during nanoparticle formation such as loss of fluorescence and size polydispersity. Fluorescence intensity of the nanoparticles was found to be size dependent, and was optimized for differently sized nanoparticles. The resulting NIR nanoparticles were also found to be more fluorescent and highly photostable compared to the free dye in solution, showing their potential as biomaterials for in vivo fluorescence imaging.

  • RESEARCH ARTICLE: Analysis of Photoexcitation Energy Dependence in the Photoluminescence of Firefly Luciferin

    RESEARCH ARTICLE: Analysis of Photoexcitation Energy Dependence in the Photoluminescence of Firefly Luciferin

    The whole pathways for photoluminescence, which include absorption, relaxation and emission, of firefly luciferin in aqueous solutions of different pH values with different photoexcitation energies were theoretically investigated by considering protonation/deprotonation. It is experimentally known that the color of fluorescence changes from green to red with a decrease in the photoexcitation energy. We confirmed with the theoretical analysis that the peak energy shift in the fluorescence spectra with varying photoenergies is due to a change in photoluminescence pathway. When the photoexcitation energy is decreased, the red emission from a monoanion form of firefly luciferin with carboxylate and phenolate groups and N-protonated thiazoline ring occurs irrespective of the pH values. However, because the species abundant in the solution and those excited by the photon depend on the solution pH, the pathway leading to the monoanion form changes with the solution pH.

  • RESEARCH ARTICLE: Effect of Laser Phototherapy on Enzymatic Activity of Salivary Glands of Hamsters Treated with 5-Fluorouracil

    RESEARCH ARTICLE: Effect of Laser Phototherapy on Enzymatic Activity of Salivary Glands of Hamsters Treated with 5-Fluorouracil

    The chemotherapeutic agent 5-Fluorouracil (5-FU) can induce salivary gland hypofunction (SGH); however, previous studies did not reach final conclusions on the influence of this drug on glandular tissue. Thus, the aim of this study was to investigate the effect of 5-FU on submandibular (SMs) and sublingual glands (SLs), as well as, the effect of laser phototherapy (LPT) on SGH induced by 5-FU. Eighty-five hamsters were divided into three groups: control (C), chemotherapy (CT) and laser (L), and the SGH was induced by two injections of 5-FU in groups CT and L. The irradiation was performed using a diode (λ780 nm/20 mW/5 J cm−2/0.2 J and 10 s per point/spot size of 0.04 cm2) and applied daily. On the euthanasia day, SMs and SLs were removed and biochemical analyses were carried out. The lactate dehydrogenase activity was increased in group CT when compared with group C for SLs and SMs (P < 0.05). In addition, the peroxidase and catalase activities were increased and superoxide dismutase was decreased by 5-FU (P < 0.05). However, LPT appears to be a protective mechanism against oxidative stress, tending to alter the activity of these antioxidant enzymes, suggesting LPT as a promising therapy to modulate the 5-FU harmful effect.

  • RESEARCH ARTICLE: Practical Labeling Methodology for Choline-Derived Lipids and Applications in Live Cell Fluorescence Imaging

    RESEARCH ARTICLE: Practical Labeling Methodology for Choline-Derived Lipids and Applications in Live Cell Fluorescence Imaging

    Lipids of the plasma membrane participate in a variety of biological processes, and methods to probe their function and cellular location are essential to understanding biochemical mechanisms. Previous reports have established that phosphocholine-containing lipids can be labeled by alkyne groups through metabolic incorporation. Herein, we have tested alkyne, azide and ketone-containing derivatives of choline as metabolic labels of choline-containing lipids in cells. We also show that 17-octadecynoic acid can be used as a complementary metabolic label for lipid acyl chains. We provide methods for the synthesis of cyanine-based dyes that are reactive with alkyne, azide and ketone metabolic labels. Using an improved method for fluorophore conjugation to azide or alkyne-modified lipids by Cu(I)-catalyzed azide-alkyne cycloaddition (CuAAC), we apply this methodology in cells. Lipid-labeled cell membranes were then interrogated using flow cytometry and fluorescence microscopy. Furthermore, we explored the utility of this labeling strategy for use in live cell experiments. We demonstrate measurements of lipid dynamics (lateral mobility) by fluorescence photobleaching recovery (FPR). In addition, we show that adhesion of cells to specific surfaces can be accomplished by chemically linking membrane lipids to a functionalized surface. The strategies described provide robust methods for introducing bioorthogonal labels into native lipids.

  • RESEARH ARTICLE: Inactivation of Bacteriophage Infecting Bacteroides Strain GB124 Using UV-B Radiation

    RESEARH ARTICLE: Inactivation of Bacteriophage Infecting Bacteroides Strain GB124 Using UV-B Radiation

    Ultraviolet-B radiation (280–320 nm) has long been associated with the inactivation of microorganisms in the natural environment. Determination of the environmental inactivation kinetics of specific indicator organisms [used as tools in the field of microbial source tracking (MST)] is fundamental to their successful deployment, particularly in geographic regions subject to high levels of solar radiation. Phage infecting Bacteroides fragilis host strain GB124 (B124 phage) have been demonstrated to be highly specific indicators of human fecal contamination, but to date, little is known about their susceptibility to UV-B radiation. Therefore, B124 phage (n = 7) isolated from municipal wastewater effluent, were irradiated in a controlled laboratory environment using UV-B collimated beam experiments. All B124 phage suspensions possessed highly similar first order log-linear inactivation profiles and the mean fluence required to inactivate phage by 4 − log10 was 320 mJ cm−2. These findings suggest that phage infecting GB124 are likely to be inactivated when exposed to the levels of UV-B solar radiation experienced in a variety of environmental settings. As such, this may limit the utility of such methods for determining more remote inputs of fecal contamination in areas subject to high levels of solar radiation.

  • RESEARCH ARTICLE: The Number of Accumulated Photons and the Quality of Stimulated Emission Depletion Lifetime Images
  • RESEARCH NOTE: Preparation and Characterization of Uniform Near IR Polystyrene Nanoparticles
  • RESEARCH ARTICLE: Analysis of Photoexcitation Energy Dependence in the Photoluminescence of Firefly Luciferin
  • RESEARCH ARTICLE: Effect of Laser Phototherapy on Enzymatic Activity of Salivary Glands of Hamsters Treated with 5-Fluorouracil
  • RESEARCH ARTICLE: Practical Labeling Methodology for Choline-Derived Lipids and Applications in Live Cell Fluorescence Imaging
  • RESEARH ARTICLE: Inactivation of Bacteriophage Infecting Bacteroides Strain GB124 Using UV-B Radiation

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Volume 87

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16th International Congress on Photobiology

September 8th – 12th, 2014

Universidad Nacional de Cordoba Cordoba, Argentina

Registrations are open: http://www.photobiology2014.com.ar/

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The photoluminescence of firefly luciferin in aqueous solutions and the pathway for red emission are shown. It is experimentally known that the colour of this fluorescence changes from green to red with a decrease in the photoexcitation energy. Hiyama and co-workers (pp. 820 – 828) report the entire pathways for photoluminescence – including absorption, relaxation and emission – of firefly luciferin in aqueous solutions of different pH values with different photoexcitation energies using quantum chemistry calculations. This theoretical study revealed that the peak energy shift in the fluorescence spectra with varying photoenergies is due to a change in photoluminescence pathway. When the photoexcitation energy is decreased, the red emission occurs, irrespective of the pH values, from a monoanion form of firefly luciferin with carboxylate and phenolate groups and N-protonated thiazoline ring. However, because the species abundant in the solution and those excited by the photon depend on pH, the pathway leading to the monoanion form changes with the solution pH. A similar analysis is possible for oxyluciferin, which has the same basic structure as firefly luciferin. In addition, the pKa obtained in this study will be useful for understanding the mechanism of firefly bioluminescence.
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