Antigenotoxicity and antimutagenicity of lycopene in HepG2 cell line evaluated by the comet assay and micronucleus test

Epidemiological studies have provided evidence that high consumption of tomatoes effectively reduces the risk of reactive oxygen species (ROS)-mediated diseases such as cancer. Tomatoes are rich sources of lycopene, a potent singlet oxygen-quenching carotenoid. In addition to its antioxidant properties, lycopene shows an array of biological effects including antimutagenic and anticarcinogenic activities. In the present study, the chemopreventive action of lycopene was examined on DNA damage and clastogenic or aneugenic effects of H2O2 and n-nitrosodiethylamine (DEN) in the metabolically competent human hepatoma cell line (HepG2 cells). Lycopene at concentrations of 10. 25, and 50 mu M, was tested under three protocols: before, simultaneously, and after treatment with the mutagen, using the comet and micronucleus assays. Lycopene significantly reduced the genotoxicity and mutagenicity of H2O2 in all of the conditions tested. For DEN, significant reductions of primary DNA damage (comet assay) were detected when the carotenoid (all of the doses) was added in the cell culture medium before or simultaneously with the mutagen. In the micronucleus test, the protective effect of lycopene was observed only when added prior to DEN treatment. In conclusion, our results suggest that lycopene is a suitable agent for preventing chemically-induced DNA and chromosome damage. (C) 2007 Elsevier Ltd. All rights reserved.

Experimental selective choriocapillaris photothrombosis using a modified indocyanine green formulation

Background: This in vivo study assessed and compared the effectiveness of an aqueous indocyanine green (ICG) formulation (R-ICG) and a lipid ICG formulation (L-ICG) in occluding the rabbit choriocapillaris, and determined the singlet oxygen quantum yields and aggregation properties of both formulations in vitro. Methods: Singlet oxygen production and aggregation were compared. The eye fundus of 30 albino rabbits was irradiated 0-15 min after dye injection using an 810 nm diode laser. Fluorescein angiography and light microscopy were used to evaluate the safety and efficacy of R-ICG and L-ICG. Results: L-ICG decreased the dimerisation constant and the tendency of ICG to form aggregates, and increased the efficiency of ICG in generating singlet oxygen (R-ICG, Phi Delta= 0.120 and L-ICG, Phi Delta= 0.210). Using a 10 mg/kg dose, choriocapillaris occlusion was achieved at a light dose of 35.8 J/cm(2) with L-ICG and 71.6 J/cm(2) with R-ICG with minimal damage to the neurosensory retina. Conclusion: Restrictions to the use of ICG in aqueous solution, low singlet oxygen quantum yields and high aggregation tendency, were overcome with L-ICG. The lower laser irradiance required to obtain choriocapillaris occlusion may suggest that L-ICG is a more potent and selective photosensitiser than R-ICG.

Propietats d'absorció bifotònica del tetrafenilporficè i el seu complex de palladi: potencials sensibilitzadors per a Teràpia Fotodinàmica bifotònica.

Estudi elaborat a partir d’una estada a la Universitat de Aarhus, Dinamarca, durant juliol 2006. En el present treball s’investiguen, per primer cop, les propietats de absorció bifotòniques del fotosensibilitzador 2,7,12,17-tetrafenilporficè (TPPo) i del seu complex de pal•ladi (II) (PdTPPo). Ambos compostos han rebut molta atenció com a possibles otosensibilitzadors per a Teràpia Fotodinàmica (TFD). S’utilitza la detecció de la fosforescència de l’oxigen singlet, centrada a 1270 nm i produïda per l’absorció de dos fotons, per quantificar la magnitud de la secció d’absorció bifotònica, "delta", dels porficèns estudiats. Els experiments se han dut a terme en el marge espectral 750-850 nm i a l’infraroig proper a 1100nm. Aquestes longituds d’ona corresponen a les zones d’absorció bifotònica en les bandes de Soret i Q. Les propietats bifotòniques obtingudes es comparen i contrasten amb les dades conegudes de la tetrafenilporfirina (TPP), isòmer estructural del tetrafenilporficè però amb més gran simetria, i es troba que en la banda de Soret (que coincideix amb la regió de la pell més transparent) els valors de delta per el TPPo i PdTPPo son aproximadament 2000 GM en el màxim, pràcticament cent vegades més grans que per la TPP. A més a més, aquestos valors son dos ordres de magnitud més grans que els obtinguts a l’irradiar a 1100 nm (bandes Q). Aquestes observacions es poden explicar mitjançant la amplificació per ressonància deguda a la presència de transicions monofotòniques ressonants en la regió de les bandes Q. Els elevats valors de "delta" obtinguts per els tetrafenilporficèns estudiats junt a les principals característiques que aquestos presenten (elevat rendiment de formació d’oxigen singlet, estabilitat química i fotoquímica, absència de citotoxicitat,...) qualifiquen al TPPo y PdTPPo com a possibles fotosensibilitzadors per a Teràpia Fotodinàmica Bifotònica.

Fotofísica de carotenóides e o papel antioxidante de b-caroteno

Carotenoid polyenes play a wide role in nature and their photophysical properties make of these pigments a focus of research in photochemistry, photobiology and photomedicine. Some aspects of the singlet and triplet states and, their interaction with molecular and singlet oxygen and free radicals are briefly reviewed in this article.

Quimiluminescência orgânica: alguns experimentos de demonstração para a sala de aula

The generation of "cold light", visible to the human eye, by chemical reactions has attracted the attention of the scientific community since the beginning of this century. Besides the academic interest in the elucidation of the mechanisms of excited state formations, many chemiluminescence reactions have found widespread analytical applications. Moreover, the phenomenon of chemiluminescence can also be used as a tool in undergraduate and college teaching. In this article, we describe several known chemiluminescence demonstrations, which are suitable for school teaching. The main objective of this work is to produce didactic material in Portuguese to stimulate Brazilian secondary and high school teachers to use these experiments in the classroom. The demonstrations include singlet oxygen emission, the luminol reaction, oscillating chemiluminescence and the peroxyoxalate reaction.

Terapia fotodinâmica: princípios, potencial de aplicação e perspectivas

Photodynamic Therapy (PDT) has been designated as a promising new modality in the treatment of cancer and other diseases since the early 1980s. It has been used with success for the treatment of a variety of tumours, and attempts are being made to extend this treatment modality to other clinical conditions (as example, the inactivation of viruses in blood and blood components). This can be partly attributed to the very attractive basic concept of PDT: the combination of a photosensitizing drug and light, which are relatively harmless by themselves but combined (in the presence of oxygen) ultimately cause more or less selective tumour destruction.

Utilização de endoperóxidos de derivados de naftaleno como fontes químicas de oxigênio singlete em sistemas biológicos

Molecular oxygen, in the first excited state (singlet oxygen, ¹O2), has a substantial reactivity towards electron-rich organic molecules, such as biological targets, including unsaturated fatty acids, proteins, RNA and DNA. Considering the complexity of biological systems and the great variety of reactive species generated by photochemistry, efforts have been devoted to develop suitable ¹O2 generators based on the thermolysis of water soluble naphthalene endoperoxides. These compounds are chemically inert and have been employed as versatile sources of ¹O2. The synthesis is based on structural modifications in position 1,4 of dimethylnaphtalene, grafting hydrophilic substituents. The correspondent endoperoxide can be generated using photochemical method, or molybdate-catalyzed disproportionation of hydrogen peroxide.

Carotenóides: propriedades, aplicações e biotransformação para formação de compostos de aroma

Carotenoids are widely distributed in nature, providing yellow, orange or red color in a great number of vegetables, microorganisms and in some animals. Carotenoids act as biological antioxidants and seem to play an important role in human health by protecting cells and tissues from the damaging effects of free radicals and singlet oxygen. Several authors describe the oxidative cleavage of carotenoids in flavor compounds as occuring through chemical or photochemical degradations or through biotechnological processes. Biotransformation of carotenoids seems to be a reasonable alternative to produce flavor compounds since these compounds are considered 'natural' ingredients. In this work we describe the properties of some carotenoids, as well as biotechnological approaches to obtain its oxyfunctionalized derivatives.

Superoxide: a two-edged sword

Superoxide (O2-) is the compound obtained when oxygen is reduced by one electron. For a molecule with an unpaired electron, O2- is surprisingly inert, its chief reaction being a dismutation in which it reacts with itself to form H2O2 and oxygen. The involvement of O2- in biological systems was first revealed by the discovery in 1969 of superoxide dismutase, an enzyme that catalyzes the dismutation of O2-. Since then it has been found that biological systems produce a bewildering variety of reactive oxidants, all but a few arising ultimately from O2-. These oxidants include O2- itself, H2O2 and alkyl peroxides, hydroxyl radical and other reactive oxidizing radicals, oxidized halogens and halamines, singlet oxygen, and peroxynitrite. These various oxidants are able to damage molecules in their environment, and are therefore very dangerous. They are thought to participate in the pathogenesis of a number of common diseases, including among others malignancy, by their ability to mutate the genome, and atherosclerosis, by their capacity for oxidizing lipoproteins. Their properties are put to good use, however, in host defense, where they serve as microbicidal and parasiticidal agents, and in biological signalling, where their liberation in small quantities results in redox-mediated changes in the functions of enzymes and other proteins

Photochemistry and photobiology of actinic erythema: defensive and reparative cutaneous mechanisms

Sunlight is part of our everyday life and most people accept it as beneficial to our health. With the advance of our knowledge in cutaneous photochemistry, photobiology and photomedicine over the past four decades, the terrestrial solar radiation has become a concern of dermatologists and is considered to be a major damaging environmental factor for our skin. Most photobiological effects (e.g., sunburn, suntanning, local and systemic immunosuppression, photoaging or dermatoheliosis, skin cancer and precancer, etc.) are attributed to ultraviolet radiation (UVR) and more particularly to UVB radiation (290-320 nm). UVA radiation (320-400 nm) also plays an important role in the induction of erythema by the photosensitized generation of reactive oxygen species (singlet oxygen (1O2), superoxide (O2.-) and hydroxyl radicals (.OH)) that damage DNA and cellular membranes, and promote carcinogenesis and the changes associated with photoaging. Therefore, research efforts have been directed at a better photochemical and photobiological understanding of the so-called sunburn reaction, actinic or solar erythema. To survive the insults of actinic damage, the skin appears to have different intrinsic defensive mechanisms, among which antioxidants (enzymatic and non-enzymatic systems) play a pivotal role. In this paper, we will review the basic aspects of the action of UVR on the skin: a) photochemical reactions resulting from photon absorption by endogenous chromophores; b) the lipid peroxidation phenomenon, and c) intrinsic defensive cutaneous mechanisms (antioxidant systems). The last section will cover the inflammatory response including mediator release after cutaneous UVR exposure and adhesion molecule expression

The photodynamic and non-photodynamic actions of porphyrins

Porphyrias are a family of inherited diseases, each associated with a partial defect in one of the enzymes of the heme biosynthetic pathway. In six of the eight porphyrias described, the main clinical manifestation is skin photosensitivity brought about by the action of light on porphyrins, which are deposited in the upper epidermal layer of the skin. Porphyrins absorb light energy intensively in the UV region, and to a lesser extent in the long visible bands, resulting in transitions to excited electronic states. The excited porphyrin may react directly with biological structures (type I reactions) or with molecular oxygen, generating excited singlet oxygen (type II reactions). Besides this well-known photodynamic action of porphyrins, a novel light-independent effect of porphyrins has been described. Irradiation of enzymes in the presence of porphyrins mainly induces type I reactions, although type II reactions could also occur, further increasing the direct non-photodynamic effect of porphyrins on proteins and macromolecules. Conformational changes of protein structure are induced by porphyrins in the dark or under UV light, resulting in reduced enzyme activity and increased proteolytic susceptibility. The effect of porphyrins depends not only on their physico-chemical properties but also on the specific site on the protein on which they act. Porphyrin action alters the functionality of the enzymes of the heme biosynthetic pathway exacerbating the metabolic deficiencies in porphyrias. Light energy absorption by porphyrins results in the generation of oxygen reactive species, overcoming the protective cellular mechanisms and leading to molecular, cell and tissue damage, thus amplifying the porphyric picture.

Photodynamic DNA damage induced by phycocyanin and its repair in Saccharomyces cerevisiae

In the present study, we analyzed DNA damage induced by phycocyanin (PHY) in the presence of visible light (VL) using a set of repair endonucleases purified from Escherichia coli. We demonstrated that the profile of DNA damage induced by PHY is clearly different from that induced by molecules that exert deleterious effects on DNA involving solely singlet oxygen as reactive species. Most of PHY-induced lesions are single strand breaks and, to a lesser extent, base oxidized sites, which are recognized by Nth, Nfo and Fpg enzymes. High pressure liquid chromatography coupled to electrochemical detection revealed that PHY photosensitization did not induce 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodGuo) at detectable levels. DNA repair after PHY photosensitization was also investigated. Plasmid DNA damaged by PHY photosensitization was used to transform a series of Saccharomyces cerevisiae DNA repair mutants. The results revealed that plasmid survival was greatly reduced in rad14 mutants, while the ogg1 mutation did not modify the plasmid survival when compared to that in the wild type. Furthermore, plasmid survival in the ogg1 rad14 double mutant was not different from that in the rad14 single mutant. The results reported here indicate that lethal lesions induced by PHY plus VL are repaired differently by prokaryotic and eukaryotic cells. Morever, nucleotide excision repair seems to play a major role in the recognition and repair of these lesions in Saccharomyces cerevisiae.

Photodynamic antimicrobial chemotherapy (PACT) for the treatment of malaria, leishmaniasis and trypanosomiasis

A photodynamic effect occurs when photosensitiser molecules absorb light and dissipate the absorbed energy by transferring it to biological acceptors (usually oxygen), generating an excess of reactive species that are able to force cells into death pathways. Several tropical diseases present physiopathological aspects that are accessible to the application of a photosensitiser and local illumination. In addition, disease may be transmitted through infected blood donations, and many of the aetiological agents associated with tropical diseases have been shown to be susceptible to the photodynamic approach. However, there has been no systematic investigation of the application of photoantimicrobial agents in the various presentations, whether to human disease or to the disinfection of blood products or even as photo-insecticides. We aim in this review to report the advances in the photoantimicrobial approach that are beneficial to the field of anti-parasite therapy and also have the potential to facilitate the development of low-cost/high-efficiency protocols for underserved populations.

Investigation of the Photodynamic Effects of Curcumin Against Candida albicans

This study describes the association of curcumin with light emitting diode (LED) for the inactivation of Candida albicans. Suspensions of Candida were treated with nine curcumin concentrations and exposed to LED at different fluences. The protocol that showed the best outcomes for Candida inactivation was selected to evaluate the effect of the preirradiation time (PIT) on photodynamic therapy (PDT) effectiveness, the uptake of curcumin by C. albicans cells and the possible involvement of singlet oxygen in the photodynamic action. Curcumin-mediated PDT was also assessed against biofilms. In addition to the microbiological experiments, similar protocols were tested on a macrophage cell line and the effect was evaluated by Methyltetrazolium assay (MTT) and SEM analysis. The optical properties of curcumin were investigated as a function of illumination fluence. When compared with the control group, a statistically significant reduction in C. albicans viability was observed after PDT (P < 0.05), for both planktonic and biofilm cultures. Photodynamic effect was greatly increased with the presence of curcumin in the surrounding media and the PIT of 20 min improved PDT effectiveness against biofilms. Although PDT was phototoxic to macrophages, the therapy was more effective in inactivating the yeast cell than the defense cell. The spectral changes showed a high photobleaching rate of curcumin.

Influence of pH on the phototransformation process of PhotogemA (R)

PhotogemA (R) is a hematoporphyrin derivative that has been used as a photosensitizer in experimental and clinical Photodynamic Therapy (PDT) in Brazil. Photosensitizers are degraded under illumination. This process, usually called photobleaching, can be monitored by decreasing in fluorescence intensities and includes the following photoprocesses: photodegradation, phototransformation, and photorelocalization. Photobleaching of hematoporphyrin-type sensitizers during illumination in aqueous solution is related not only to photodegradation but is also followed by the formation of photoproducts with a new fluorescence band at around 640-650 nm and with increased light absorption in the red spectral region at 640 nm. In this study, the influence of pH on the phototransformation process was investigated. PhotogemA (R) solutions, 40 mu g/ml, were irradiated at 514 nm with intensity of 100 mW/cm(2) for 20 min with different pH environments. The controls were performed with the samples in the absence of light. The PhotogemA (R) photodegradation is dependent on the pH. The behavior of photodegradation and photoproducts formation (monitored at 640 nm) is distinct and depends on the photosensitizer concentration. The processes of degradation and photoproducts formation were monitored with Photogemin the concentration of 40 mu g/mL since that demonstrated the best visualization of both processes. While below pH 5 the photodegradation occurred, there was no detectable presence of photoproducts. The increase of pH led to increase of photoproducts formation rate with photodegradation reaching the highest value at pH 10. The increase of photoproducts formation and instability of PhotogemA (R) from pH 6 to pH 10 are in agreement with the desired properties of an ideal photosensitizer since there are significant differences in pH between normal (7.0 < pH < 8.6) and tumor (5.8 < pH < 7.9) tissues. It is important to know the effect of pH in the process of phototransformation (degradation and photoproduct formation) of the molecule since low pH values promotes increase in the proportion of aggregates species in solution and high pH values promotes increase in the proportion of monomeric species. There must be an ideal pH interval which favors the phototransformation process that is correlated with the singlet oxygen formation responsible by the photodynamic effect. These differences in pH between normal and tumor cells can explain the presence of photosensitizers in target tumor cells, making PDT a selective therapy.